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Year 1 Semester 1 > Biochemistry > Flashcards

Biochemistry Flashcards

1
Q

What can be made up by protein?

A

-Cytoskeleton
-Enzymes
-Cell receptors
-Toxins
-Antibodies
-Hair
-Fingernails
-Histones
-Transporters
-Ribosomal proteins

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2
Q

How many different proteins are present in humans?

A

20000

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3
Q

What are the four levels of polypeptide structure?

A

-Primary
-Secondary
-Tertiary
-Quaternary

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4
Q

How are peptide bonds formed?

A

Through Condensation reactions between the carboxylic acid of one AA and the amino group of another.

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5
Q

What is the length of a C=N Peptide bond?

A

1.32 Angstrom

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6
Q

What kind of bond holds together the polypeptide bond?

A

Covalent bond

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7
Q

In what ways are proteins chiral?

A

-They are made up of L-amino acids
-Secondary structures have handedness (alpha helices are right-handed, and beta strands twist)
-Their oligomeric states can be handed

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8
Q

What is the primary protein structure?

A

The individual linear sequence of amino acids that make up a protein.

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9
Q

How many proteinogenic amino acids are there?

A

21 (but 1 isn’t produced in the body)

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10
Q

How can we write amino acids?

A

-Use full name without capital letter (eg arginine)
-Three letter code (with position in the chain eg Arg79)
-One letter code used in sequence (egR234)

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11
Q

What is each amino acid in a chain called?

A

Residue

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12
Q

What can then polypeptide chain structure rotate around?

A

The alpha carbon

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13
Q

What does phi rotation refer to in the primary structure of a polypeptide?

A

Rotation around the C’-N-aC-C’

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14
Q

What does psi rotation refer to in the primary structure of a polypeptide?

A

Rotation around N-aC-C’-N

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15
Q

What does omega rotation refer to in the primary structure of a polypeptide?

A

aC- C-N-aC

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16
Q

How many degrees is the rotation in a polypeptide chain?

A

either 180 (trans) or 0 (cis)

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17
Q

Describe the alpha-helix secondary structure

A

-Right handed helix
-100 degrees repeat ~3.6 residues per turn
-Pitch 5.4 Armstrong
-Satisfies H-bonding of backbone
-Side chains point outwards
-Prefers Ala, Glu, Leu, Met, Glu, Ile, Lys, Phe Trp (no Pro)
-Average 11 residues

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18
Q

Describe the beta-strand secondary structure

A

-2 residue repeat eg every second side chain points in the same direction.
-H-bonds are shared with other strands
-Both have 2 residue repeats of 7 Amstrong
-Prefer Ile, Tyr, Val
-2-15 strands, average 6 strands
-Parallel is less stable than anti-parallel
-Tend to have a right-handed twist

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19
Q

How much of the protein structure do Alpha Helices and Beta-strands make up?

A

Roughly 50%.

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20
Q

What are the 6 Aliphatic Amino Acids?

A

Alanine, Glycine, Isoleucine, Leucine, Proline, and Valine

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20
Q

What are the 3 aromatic Amino acids?

A

Phenylalanine, Tryptophan, and Tyrosine

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21
Q

What are the 2 acidic Amino Acids?

A

Aspartic acid and Glutamic Acid

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22
Q

What are the 3 Basic Amino acids?

A

Arginine, Histidine and Lysine

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23
Q

What are the 2 Hydroxylic Amino acids?

A

Serine and Threonine

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24
Q

What are the two Sulphur containing Amino Acids?

A

Cysteine and Methionene

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25
Q

What are the two Amidic Amino acids?

A

Asparagine and Glutamine

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26
Q

What are the 2 Amino acids that are not coded for by codons in the human body?

A

Selenocysteine and Hydroxyproline

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27
Q

What can a lack of Hydroxyproline result in?

A

Less stable collagen (leading to scurvy).

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28
Q

What is Alanine’s 3 letter code and 1 letter code?

A

Ala + A

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29
Q

What is Arginine’s 3 letter code and 1 letter code?

A

Arg + R

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30
Q

What is asparagine’s 3 letter code and 1 letter code?

A

Asn + N

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31
Q

What is aspartic acid’s 3 letter code and 1 letter code?

A

Asp + D

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32
Q

What is Cysteine’s 3 letter code and 1 letter code?

A

Cys + C

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33
Q

What is Glutamic acid’s 3 letter code and 1 letter code?

A

Glu + E

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34
Q

What is Glutamine’s 3 letter code and 1 letter code?

A

Gln + Q

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35
Q

What is glycine’s 3 letter code and 1 letter code?

A

Gly + G

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36
Q

What is Histidine’s 3 letter code and 1 letter code?

A

His + H

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37
Q

What is Isoleucine’s 3 letter code and 1 letter code?

A

Ile + I

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38
Q

What is Leucine’s 3 letter code and 1 letter code?

A

Leu + L

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39
Q

What is Lysine’s 3 letter code and 1 letter code?

A

Lys + K

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40
Q

What is Methionine’s 3 letter code and 1 letter code?

A

Met + M

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41
Q

What is Phenylalanine’s 3 letter code and 1 letter code?

A

Phe + F

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42
Q

What is Proline’s 3 letter code and 1 letter code?

A

Pro + P

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43
Q

What is Serine’s 3 letter code and 1 letter code?

A

Ser + S

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44
Q

What is Threonine’s 3 letter code and 1 letter code?

A

Thr + T

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45
Q

What is Tryptophan’s three letter code and 1 letter code?

A

Trp + W

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46
Q

What is Tyrosine’s 3 letter code and 1 letter code?

A

Tyr + Y

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47
Q

What is Valine’s 3 letter code and 1 letter code?

A

Val + V

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48
Q

What is Selenocysteine’s 3 letter code and 1 letter code?

A

Sec + U

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49
Q

What is Hydroxyproline’s 3 letter code and 1 letter code?

A

Hyp + O

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50
Q

How are Alpha helices and Beta-Strands formed?

A

Through Hydrogen bonding between the NH and CO of amino acids in a polypeptide sequence.

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51
Q

What does the overall structure of a protein determine?

A

The overall structure of a protein determines how it will interact with other proteins, biomolecules and small molecules and thus determines how it will function.

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52
Q

How can we determine the structure of a protein?

A

Eg -Through Xray crystallography
-AlphaFold technology

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53
Q

What is the tertiary structure of a protein?

A

The tertiary structure of a protein is its overall 3D shape. The final folded protein is usually its active form.

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54
Q

What is a domain (with regards to a protein)?

A

Proteins often fold in chunks known as domains. Domains are independent folding units of structure, typically 100-200 AA long.

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55
Q

What is the quaternary structure of a protein?

A

How multiple protein chains bind together (homo/hetero interactions), or to small molecules/metals/cofactors.

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56
Q

Describe how myoglobin and haemoglobin bind to the haem cofactor.

A

-Porphyrin ring with iron associated
-Globin protein coordinates iron axially
-Free coordination binds gases

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57
Q

What is Allostery?

A

Binding at one site affects the binding at another site.

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58
Q

What affects the oxygen affinity of haemoglobin?

A

-Amount of oxygen bound (Allostery)
-Chloride ions
-Bisphosphoglycerate
-Protonation state of haemoglobin
-CO2 binding

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59
Q

What happens if a protein that is developmentally essential is missing or non-functioning?

A

Death

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60
Q

What happens if a protein is missing or non-functioning

A

Disease

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61
Q

Describe cystic fibrosis.

A

-Caused by mis-sense mutations (such as G542X, W1282X, R553X) in the CFTR (Cystic fibrosis transmembrane regulator) gene.
-Leads to absence or dysfunctional ATP-gated Anion channel that regulates extracellular Cl- in the epithelial tissues.
-Causes thick mucus, Poor secretion of digestive enzymes from pancreas, and infertility.

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62
Q

Describe phenylketonuria.

A

-Caused by the loss or dysfunctional phenylalanine hydroxylate enzyme
-This would normally metabolise phenylalanine to tyrosine (phenylalanine in excess causes neurological and developmental disorders)

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63
Q

Describe Type 1 Diabetes.

A

-Autoimmune disease, attacking the pancreatic beta cells (which produce the insulin protein).
-Leads to hypoglycaemia, sweats and seizures, nerve damage, loss of limbs.

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64
Q

Describe insulin.

A

-Produced as 110aa preproinsulin in beta cells in pancreas
-Folds in endoplasmic reticulum and processed by proteases to give a protein of 51 amino acids
-Stored at zinc-coordinated hexers in crystals in the pancreas
-Acts as a monomer on insulin receptors on cells.

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65
Q

Give examples of important non-globular proteins.

A

-Microfilaments
-Intermediate filaments
-Keratin
-Vimentin
-Alpha internexin/nestin
-Lamin
-Microtubules
-Muscle fibres
-Cilia
-Flagella
-Collagen
-Silk

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66
Q

Describe protein fibres.

A

-Globular proteins that assemble into fibrous quaternary arrangements
Tend to be
-Strong
-Dynamic
-Able to interact with other proteins and DNA

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67
Q

Describe globular proteins

A

Well folded, have distinct secondary and tertiary structures, approximately spherical and are most enzymes and enzyme subunits.

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68
Q

What are intrinsically disordered proteins?

A

Proteins without regular tertiary structures that remain stable and active

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69
Q

Give examples of diseases that result from protein misfolding

A

-Alzheimers (beta amyloid plaque)
-Parkinson’s (alpha-synuclein plaques)
-ALS (non degraded proteins in motor neurones)
-CJD (Prions aggregates in the brain)

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70
Q

What is protein aggregation?

A

Misfiling to form predominantly beta-sheet structures.

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71
Q

What are amyloids?

A

Distinct cross-beta-sheet arrangements, found across kingdoms and in different diseases.

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72
Q

Give some key proteins found in Influenza.

A

Haemagglutinin - Facilitates entry of virus into host cells by binding to sugars on host cell surface, fusing membranes and releasing the virus into the cell.
Neuraminidase - Facilitates exit from host cell by binding to sugars on host cell surface and cleaving sialic acid sugars from cell glycoproteins. This prevents the virus sticking to cells.

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73
Q

Give examples of Antiflu drugs.

A

-Tamiflu and Relenza
-Target neuraminidase enzyme of influenza, competitively binding with the enzyme due to its similar structure to neuraminic acid.

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74
Q

Give examples of mutations in the neuraminidase gene that give resistance to tamiflu.

A

-H274Y
-R292K

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75
Q

How does Penicillin works as an antibiotic?

A

-Beta-lactam ring irreversibly interacts with active site, meaning no more crosslinks
-The cell weakens and lyses.

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76
Q

Give examples of different mechanisms for penicillin resistance.

A

-Altered binding of PBP to beta-lactums means that penicillin cannot work
-Beta lactamase enzyme breaking down penicillin
-Changes import of antibiotic
-Changes to cell wall structure

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77
Q

What are nucleoside analogue drugs?

A

-Work by interacting with DNA/RNA polymerases and reverse transcriptase enzyme
-Can block the active site
-Integrate into new RNA/DNA and block further extension of DNA/RNA molecule

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78
Q

Give examples of nucleoside analogue drugs.

A

-AZT
-Acyclovir
-Remdesvir

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79
Q

Give some examples of mechanisms of enzyme catalysis.

A

Proximity - Brings substrate molecules close together
Orientation - Orientates substrates correctly
Strain/Distortion - Binding puts strain on bond making it easier for reaction to occur
Acid-Base - Protons donated or accepted
Covalent catalysis - temporary covalent bond between enzyme and substrate

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80
Q

Give the different classes of enzymes.

A

Oxidoreductases - Oxidation/Reduction
Transferases - Transfer of a functional group from one substrate to another
Hydrolases - Formation of two products from on substrate by hydrolysis
Lyases - Non-hydrolytic addition/removal of groups
Isomerases - Intramolecular rearrangement
Ligases - Join two molecules together, synthesis of a new bond (requires ATP)
Translocases - Movement of ions or molecules across membranes

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81
Q

What is the turnover number? (Kcat)

A

The number of substrate molecules converted to product by 1 enzyme in 1 second.

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82
Q

Give the classifications of enzyme inhibitors.

A

-Irreversible
-Reversible

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83
Q

What types of reversible enzyme inhibitors are there?

A

-Competitive
-Non competitive
-Uncompetitive

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84
Q

Describe Irreversible enzyme inhibitorsn.

A

-Bind irreversibly to enzyme, usually via a covalent bond
-Bond to an amino acid side chain at or near active site (commonly Ser or Cys side chains)
-Permanently binds, inactivating the enzyme through by preventing substrate binding

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85
Q

Describe competitive enzyme inhibitors.

A

-Compete with substrate for access to active site, often with a similar structure to substrate
-Can be overcome by increasing [S] until it outcompetes the inhibitor
-Most useful therapeutic agents

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86
Q

How does Vmax and Km change when competitive inhibitors are added?

A

-Km increases
-Vmax stays the same

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87
Q

Describe Non-competitive enzyme inhibitors

A

-Inhibitor binds away from the active site, modifying reaction rate
-Still binds substrate with same affinity
-Influences capacity to catalyse reaction

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88
Q

How does Vmax and Km change when Noncompetitive inhibitors are added?

A

-Km unaltered
-Vmax decreases

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89
Q

Describe Uncompetitive enzyme inhibitors

A

-Occurs with multisubstrate reactions
-Binds only to ES complex
-Km decreases
-Vmax decreases.

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90
Q

How will pH affect enzyme activity?

A

-3D structure of enzyme may change
-Groups involved in binding substrate or catalysis change through :
-Protonation/deprotonisation of side chains

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91
Q

How will temperature affect enzyme activity?

A

Can change
-diffusion rate (likelihood of forming ES complex)
-Flexibility of active site
-Stability of enzyme (denatures at high temp)
-May affect cold-active or thermostable enzymes.

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92
Q

How can molecular crowding affect enzyme activity

A

-The cell is very crowded
-Large proteins diffuse slower than small substrates/products
-In pathways with mismatched enzyme rates cells can ioncrease the amount of the slower enzyme
-In pathways with volatile intermediates cells can spatially trap them.

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93
Q

Give the steps of separating proteins through chromatography.

A

Step 1 Isolate the protein through centrifugation (if bacteria or blood) or Homogenisation (if from tissues, organs or plants)
Step 2 Prepare soluble proteins from cells by centrifuging til pellets are formed, repeating this until the correct protein supernatant is formed
Step 3 Protein isolation through either Thin layer Chromatography or Column Chromatography.

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93
Q

What are the two main methods of separating proteins?

A

Chromotography and Gel electrophoresis

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94
Q

What are the principles of chromatography

A

-Sample of interest has differential interaction strength between mobile and stationary phase
-In biological chromatography
-Stationary phase is often some kind of polymer bead
-Mobile phase is a biologically compatible buffer system specific to protein/experiment being performed.

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95
Q

What can we use to detect the eluate following a chromatogram?

A

-Measure absorbance at 280nm
-Use UV detectors
-Ion exchange
-Size exlusion
-Hydrophobic interactions
-Reversed phase

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96
Q

Describe size exclusion chromatography.

A

-AKA Gel filtration
-Separates mixtures based on hydrodynamic radius
-Large particles cannot enter gel and are excluded, meaning they have less volume to traverse and elute sooner, whereas smaller particles can enter gel and have more volume to traverse, so elute later.

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97
Q

Describe Ion exchange chromatography.

A

-Separates proteins with differences in surface charge
-Proteins elute sooner/later to a negative or positive stationary phase based on charge.

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98
Q

Describe separation of proteins based on hydrophobicity.

A

-Separation of mixtures based on surface hydrophobicity
-Media modified with hydrophobic alkyl/aryl groups
-High salt conc favours interatction with media.

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99
Q

Describe affinity chromatography.

A

-Stationary phase functionalised with chemical/protein that binds protein of interest.

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100
Q

Describe Immobilised metal affinity chromatography.

A

-Requires recombinent protein expression, with tag added to protein coded gene
-His-tag is 6-10 histidine residues added to protein usually at N or C terminus.
-Histidine has an imidazole side chain that binds to metals like nickel and cobalt, producing very pure protein quickly.

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101
Q

Define Electrophoresis.

A

Movement of dispersed particles relative to a fluid under the influence of a spatially uniform electric field.

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102
Q

Describe gel electrophoresis.

A

Polymer gels act as molecular sieves, with pore size depending on concentration of gelling agent.

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103
Q

Give the components of Polymer gels used in electrophoresis.

A

Agarose - A linear polymer extracted from red seaweed, used to separate nucleic acids/protein complexes.
Acrylamide - An organic compound with multiple functional groups, crosslinked using a catalyst, used to separate proteins/nucleic acids.
Buffering - A liquid phase buffer to reduce current induced pH changes and minimise heating.

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104
Q

What factors affect gel electrophoresis.

A

-Net charge of molecule
-Size of molecule
-Electric field strength
-Properties of gel
-pH, counter ions and salts found in running buffer
-Temperature

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105
Q

Give different variants of Polyacrylamide Gel electrophoresis.

A

-Native PAGE
-Blue Native PAGE
-SDS PAGE

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106
Q

Describe Native PAGE.

A

Polyacrylamide Gel electrophoresis
-Used to separate acidic proteins
-Separation by charge/size
-Samples added to buffer with dye/glycine/buffer
-Gels have no denaturing agent.
-Good for looking at protein complexes and post translational modifications.

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107
Q

How does Blue Native PAGE differ from Native PAGE?

A

The addition of Coomassie blue, providing additional charge on proteins

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108
Q

Describe SDS-PAGE

A

-Protein denatured with heat, and SDS is added to the gel
-Formation of mixed protein:SDS Micelles
-Charge of protein masked
-Separates by mass
-Can run DNA on PAGE
-Consists of a ph6.5 stacking gel and 8.8 resolvingf gel, and glycine/chloride counter ions to buffer.
-Also contains TRIS buffer (usually pH 8.3) and SDS to ensure protein denatures.

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109
Q

Describe Agarose gel electrophoresis.

A

-0.5%-2% agarose in TAE/TBE buffer.
-Buffer consists of Tris (pH 8.3), Acetate/borate counterion, EDTA chelating agent
-Separation of molecules occurs 100-500nm
-Gives a separation range of 50bp-20kbp

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110
Q

How do we visualise molecules in gels (after electrophoresis).

A

-DNA and most proteins do not have intrinsic colour so need to be stained/dyed eg with
-Intercalating dyes: ethidium bromide
-Minor groove binders: Hoescht, DAPI
-Cyanine dues: SYBR

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110
Q

Give different dyes used in gel electrophoresis and their sensitivites.

A

-Fluor Orange 10ng
-Coomassie blue 5ng
-Silver 0.5ng
-Spyro 0.25ng

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111
Q

Give the equation to find the absorptivity of a substance.

A

Absorptivity = Molar attenuation coefficient x optical path length x concentration of attenuating species

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112
Q

What biological processes are carbohydrates involved in?

A

-Storage and transport of energy
-Cell-cell communication/adhesion
-Host-pathogen and host-symbiont reaction
-Structural components of cells
-Components of DNA and RNA

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112
Q

What are the two simplest carbohydrates that occur widely in nature containing 3 carbons?

A

Glyceraldehyde and Dihydroxyacetone

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113
Q

Why do sugar rings form in Aldehyde/Aldose carbohydrates?

A

The alcohol group attacks the aldehyde group.

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114
Q

Why do sugar rings form in Ketone/Ketose carbohydrates?

A

The alcohol group attacks the ketone group

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115
Q

Why are sugar rings not flat?

A

Because of the tetrahedral geometry of carbon atoms.

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116
Q

What are the two orientations that substituents on ring C atoms can have?

A

Axial (a) - pointing up or down
Equitorial (e) - same plane as ring

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117
Q

Describe how axial atoms are arranged on adjacent carbons

A

They point in opposite directions as if they did they would physically clash.

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118
Q

Describe ring sugar anomer formation.

A

The formation of a ring generates an additional asymmetric carbon at position 1. The stereochemical isomer at C1 is not a new sugar but is an ⍺ or β form of the same sugar, known as anomers.

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119
Q

How do you distinguish ⍺-anomers and β-anomers?

A

⍺-anomer - C1 and C5 have different stereochemistry
β-anomer - C1 and C5 have the same stereochemistry

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120
Q

How is the designation of D or L sugars determined?

A

-Determined by the configuration of the aymmetric carbon furthest from the aldehyde or keto group.

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121
Q

What are epimers?

A

Carbohydrates with a different configuration of a certain group (eg OH) at a certain asymmetric carbon.

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122
Q

Name the different forms of saccharide chains (and their lengths).

A

-Disaccharides (2 Monosaccs)
-Oligosaccharides (2-10 Monosaccs)
-Polysaccharides (>10 Monosaccs)

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123
Q

What is the difference in ⍺Glycosidic and βGlycosidic bonds?

A

-The C1 of one sugar and C5 of another is in opposite configuration in ⍺glycosidic bonds
-The C1 of one sugar and C5 of another is in equitorial configuration in βglycodisic bonds.

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123
Q

Describe how disaccharides can be used for energy transport (and give examples)

A

Disaccharides are hydrolysed by maltase, lactase and sucrase enzymes on surface of small intestine to be used as an energy source.
-Maltose (major product generated from starch digestion)
-Lactose (Milk sugar)
-Sucrose (Transport form of carbohydrate from plants)

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124
Q

Describe how polysaccharides can be used for energy transport (and give examples)

A

Starch (found in plants and act as a major human food source) and Glycogen (found in animal liver and muscle cells) acts as important energy storage. Both are polysaccharides of D-glucose.

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125
Q

Describe the structure of Starch and Glycogen.

A

-Backbone of D-glucose molecules linked by ⍺1,4glycosidic bonds
-Branches linked to backbone via ⍺1,6 glycosidic bonds
-Reducing end is the terminal sugar in which C1 is unattached (ring can open and reducing aldehyde can form)
-Non-reducing end is the terminal sugar in which C1 is involved in a glycosidic bond (preventing it from opening)

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126
Q

What are reducing sugars?

A

A reducing sugar is any sugar that is capable of acting as a reducing agent because it has a free aldehyde or ketone group.

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127
Q

Why are ⍺-glucans good storage molecules?

A

-The ⍺1,4-linkage is kinked
-Causing the polysaccharide to twist into a helical structure that is significantly more compact

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128
Q

What is peptidoglycan, and where is it typically found?

A

A structural carbohydrate that acts as a main component of the bacterial cell wall.

129
Q

What is the peptidoglycan polysaccharide chain made up of?

A

N-acetylglucosamine (GlcNAc) and N-acetylmuramic acid (MurNAc).

130
Q

Describe Nitrogen-linked glycoproteins.

A

-Joined to the amide of asparagines
-Usually on secreted proteins only
-N-glycosylation plays role in protein folding, stability and cell recognition.

131
Q

Describe Oxygen-linked proteins.

A

-Joined to hydroxyl of serine or threonine
-Common addition is GlcNAc
-Cytoplasmic
-Thought to be reciprocal to phosphorylation

132
Q

What are Proteoglycans main functions?

A

Produced by virtually all mammalian cells, acting as
-Joint lubricants
-Structural components of Extracellular Matrix
-Mediate adhesion of cells to ECM
-Bind factors that stimulate cell proliferation

133
Q

What make up proteoglycans?

A

-Made up of disaccharide repeats of amino sugar and uronic acid sugar.
-Often heavily sulfated and bind to lots of water

134
Q

What enzymes synthesise complex carbohydrates, and how?

A

GLYCOSYLTRANSFERASES transfer sugars from activated nucleotides (eg UDP) onto other molecules (eg proteins, lipids and other sugars)

135
Q

Describe the different additions to the antigens found in blood groups.

A

Blood group A - Has GalNAc
Blood group B - has Gal
Blood group AB - both GalNAc and Gal
Blood group O - Has neither

136
Q

Where is almost all organic phosphorous present in biological systems derived from?

A

Phosphate or Phosphoric acid

137
Q

What insoluble phosphate-containing mineral is found in the skeletal system?

A

Hydoxylapatite - Ca5(PO4)3(OH)

138
Q

What is an Anhydride?

A

A molecule formed by the condensation of two acid molecules

139
Q

What bonds are present in ATP, that make it highly exergenic?

A

Phosphoanhydride bonds

140
Q

At what pKa does
-Phosphoric acid
-Dihydrogen phosphate
-Hydrogen phosphate
Lose/Gain a proton

A

-2.1
-7.2
-12.7

141
Q

What is the cause for the high free energy of ATP hydrolysis?

A

-Electrostatic repulsion (phosphates are negative so repel each other)
-Resonance stabilisation (hydrolysing leads to more delocalisation of electrons in phosphates, making them more stable)
-Hydration of the products (When hydrolysed there is a greater chance for water to interact with products, making it more stable)

142
Q

What is Chargaff’s rules?

A

Adenine binds with Thymine and Cytosine binds with Guanine

143
Q

What experiment did Franklin and Wilkins perform?

A

Utilising X-ray fibre diffraction to deduce the structure of DNA.

144
Q

What did Franklin and Wilkins dicover?

A

-DNA is helical, with Meridian angle 60°
-Regular repeating units 34 Å per turn, 10 bp per turn of helix
-Helical diameter of 20Å

145
Q

Describe the sugar phosphate backbone according to the Watson and Crick model?

A

-Has a hydrophobic effect, with hydrophobic bases inside and charged backbone outside
-Has base pair stacking, with Van der Waals forces
-Negatively charged
-Targeted for non-specific DNA binding.

146
Q

Describe the differences in Major and Minor grooves of DNA

A

Major groove - 22Å wide information-rich, sequence specific DNA binding and can read the sequence without unwinding.
Minor groove - 12Å wide information-poor, less frequently used for sequence recognition, binding typically alters DNA architecture

147
Q

What are the 3 different DNA models suggested by Watson and Crick?

A

A-DNA, B-DNA, and Z-DNA

148
Q

Describe A-DNA structure.

A

-Right handed
-2.3Å per base
-25.3Å pitch
-19° base pair tilt
-2.6nm diameter

149
Q

Describe B-DNA structure

A

-Right handed
-3.4Å per base
-35.4Å pitch
-1° base pair tilt
-2.4nm diameter

150
Q

Describe Z-DNA structure

A

-Left handed
-3.8Å per base
-45.6Å pitch
-9° base pair tilt
-1.8nm diameter

151
Q

What did Kornberg discover about DNA replication.

A

-Requires dNTP precursers
-Requires a primer to begin synthesis
-Chain elongation requires 3’-OH on growing chain
-Proceeds in 5’-3’ direction
-Mistakes are corrected by 3’-5’ exonuclease enzymes

152
Q

Describe the unwinding of DNA during replication

A

-DNAB helicase, recruited by DnaA
-loaded around ssDNA
-ATPase-dependent translocation
-Strand exclusion model.

-Single stranded binding protein is loaded onto ssDNA (ssDNA wraps around tetramers) to prevent secondary structure formation.

153
Q

Describe the pre-priming complex found during DNA replication.

A

-Initiated as DnaA assembly stimulated unwinding of AT-rich array and recruites helicase (DnaB)
-DnaC is a loading factor that complexes with the C-terminus of DnaB. Following helicase closure around ssDNA, DnaC hydrolyses ATP and dissociates
-DnaG (primase) synthesises short strands of RNA known as oligonucleotides during DNA replication

154
Q

What is DnaG enzyme?

A

Primase - synthesises RNA primer and is recruited by DnaB (helicase)

155
Q

What do Topoisomerase enzymes do?

A

Relax supercoiled DNA

156
Q

How does Topoisomerase I work?

A

Catalyses relaxation of supercoiled DNA by
-Cleaving of one strand, passing the cut end under the other strand, and resealing the break
UNTANGLING

157
Q

How does Topoisomerase II work?

A

Catalyses untangling of DNA duplexes
-Cleaving both strands, pass separate duplex molecule through break, then religases the break.
UNTANGLING THE DNA

158
Q

Describe the DNA polymerase III core.

A

Palm (where catalytic activity occurs)
Fingers (Domain that grabs DNA)
Thumb (grabs newly synthesised DNA and keeps it associated with enzyme).
-Has an exonuclease domain
-βClamp increases processivity of enzyme

159
Q

What are Okazaki fragments?

A

Okazaki fragments are short sections of DNA formed at the time of discontinuous synthesis of the lagging strand during replication of DNA.

160
Q

What is Dideoxy sequencing?

A

-ddNTPs spike DNA polymerase reactions
-forming truncated products
-Allows for genetic sequencing

161
Q

Why study Gene expression in E. Coli?

A

-Bacteria are key human pathogens
-The bacterial gene expression machinery is the target for some antibiotics
-E. Coli is an important host for the production of recombinent proteins for research and industrial/medical purposes
-Understanding the process of gene expression and its control in E. Coli provides a framework for understanding it in more complex organisms.

162
Q

Define Transcription

A

Transfer of genetic information from dsDNA to ssRNA (mRNA).

163
Q

What regions of DNA are involved in transcription?

A

Promoter region - Immediately upstream of the transcribed region - transcription ‘start’ signal
Transcribed region - Bacterial mRNA can be polycistronic, allowing coordinated expression of a group of genes
Terminator - Stop signal

164
Q

Describe the E. Coli σ⁷⁰ Promoter.

A

-40-60 bp region upstream of the transcription start site (acts as a binding site for RNA polymerase)
-Contains 2 hexameric sequences located at -35 (TTGACA) and -10 (TATAAT)
-The promoter dictates where transcription starts
-Promoter strength dictates how efficiently transcription is initiated (sequences dictates strength)

165
Q

Describe E. Coli RNA polymerase (RNAP).

A

-RNA Polymerase Mg2+ dependent and multisubunit
-Core RNA Polymerase is composed of 2ɑ, 1β, 1β’, 1ω
-Core + σ factor = Holoenzyme

166
Q

Describe the difference between core vs Holoenzyme RNAP.

A

-Core RNA polymerase can catalyse the process of transcription but cannot recognise and bind to the promoter.
-σ factor binds to core to convert it to homoenzyme, and directs the recognition of promoter sequences.

167
Q

Which gene produces σ⁷⁰ factor?

A

rpoN

168
Q

When are alternative sigma factors utilised?

A

Under different environmental conditions (eg Heat shock)

169
Q

In which direction does RNA polymerase read DNA?

A

Elongation

170
Q

Where does RNA polymerase stop transcription?

A

At terminator sequences

171
Q

Give the common features of factor independent terminators.

A

-Series of 4-10 consecutive A-T base pairs
-A G+C rich region with a palindromic sequence that immediately preceds the series of A-T base pairs. This forms a hairpin loop.

172
Q

How does Rho factor terminate RNA transcription.

A

-Rho factor is composed of six identical subunits
-Rho acts as a helicase that unwinds RNA-DNA and RNA-RNA duplexes
-Rho loads onto C rich sequences (Rho utilisation site)
-RNA pol pauses at ter site
-Rho unwinds the RNA-DNA hybrid and RNA pol, mRNA and Rho are released.

173
Q

What are the two strategies for regulating transcription initiation?

A

-Negative regulatory factors called REPRESSORS
or
-Positive acting factors called ACTIVATORS

174
Q

When does a high transcription of the lac operon occur?

A

When lactose is present and glucose is absent

175
Q

Describe how the lac repressor works.

A

-The lac repressors is a 360 AA protein, forming a homotetramer, binding to the lac operator.
-Binds to a 35 bp palindrome, forming a DNA loop by binding to the Primary operator (O1) and either the O2 or O3 operators.
-When Allolactase binds to the Lac repressor its DNA binding subunits separate by 3.5A, reducing affinity (dissociating the molecule from the DNA).

176
Q

Does cAMP rise or fall when there is a large amount of glucose in the cell?

A

Glucose transport into the cell inhibits adenylate cyclase, preventing cAMP accumulation
FALLS

177
Q

Describe how Catabolite Activator Protein (CAP) protein works.

A

-cAMP binds to and activates CAP Protein.
-CAP Protein binds to DNA and helps RNA polymerase to bind to the promoter.
ACTIVATING TRANSCRIPTION.

178
Q

Give 4 features of the genetic code.

A

-It is a triplet code
-The code is non-overlapping
-The code is degenerate
-The code is Universal

179
Q

What is Transfer RNA

A

tRNAs are the adaptor molecule used for decoding the base sequence of mRNA into the amino acid sequence of proteins.

180
Q

Describe the primary structure of tRNA

A

-tRNAs are small nucleic acids of 70-90 nts
-Have a 5’ monophosphate
-Contain modified bases (Ribothymidine T, Pseudouridine ᴪ, Dihydrouridine D, Inosine I)

181
Q

Describe the secondary structure of tRNA.

A

-D loop (contains dihydrouridine)
-T loop (contains pseudouridine)
-Variable arms
-Anticodon loop
-Amino acid acceptor site

182
Q

What are tRNAs joined to amino acids known as?

A

Aminoacyl tRNAs or charged tRNAs

183
Q

Describe the reaction for the aminoacylation of tRNAs

A

-AMP is added to the carboxyl group of the amino acid to give a high energy intermediate (aminoacyl adenylate)
-Aminoacyl adenylate reacts with the appropriate uncharged tRNA to give aminoacyl tRNA and AMP.

184
Q

How do different synthetase enzymes distinguish between different tRNA molecules.

A

Using identity elements

185
Q

How do Aminoacyl tRNA synthetases proofread?

A

-Acylation site rejects amino acids that are too large
-Editing site hydrolytic cleaves amino acids that are smaller than the correct one.

186
Q

What is the Wobble hypothesis?

A

The first two bases of the codon pair with the anticodon according to the usual rules, but the base at the 5’ end of the anticodon can form non standard hdyrogen bond with the base at the 3’end of the codon.

187
Q

Give the composition of Prokaryotic ribosomes.

A

70S ribosomes contain
-a 50S subunit (31 diff L proteins, 23S rRNA and 5S rRNA)
-a 30S subunit (21 S proteins, 16S rRNA)

188
Q

Describe Initiaton during translation.

A

-The 30S subunit binds to the rbosome binding site/Shine Dalgarno sequence.
-The initiator tRNA binds to the start codon AUG.
-The 50S subunit binds to form the 70S initation complex.

189
Q

Describe the 70S initation complex between a ribosome and mRNA.

A

Consists of
-Aminoacyl site (A)
-Peptidyl site (P)
-Exit site (E)

190
Q

What is the Shine Dalgarno sequence on mRNA complementary to?

A

To the 3’ end of the 16S rRNA. This properly positions the 30S ribsomal subunit on the mRNA.

191
Q

What do Initiation factors do in translation.

A

-IF1 and IF3 bind to a free 30S subunit.
-IF2 complexed with GTP binds
-The 30S subunit attaches to a mRNA
-A charged initator tRNA can then bind and IF3 is released.
-A 50S subunit can now bind which displaces IF1 and IF2 and GTP is hydrolysed (requiring energy)

192
Q

Describe Elongation during translation.

A

-The initiator tRNA is placed in the P site.
-A new tRNA containing an amino acid enters the A site., and the anticodon is matched.
-A peptide bond is made between the two adjacent AA.
-As this bond is formed the ribosome moves 1 triplet along the mRNA, occupying the E site, and allowing an empty A site.

193
Q

What is the peptidyl transferase reaction catalysed by?

A

The 23S rRNA.

194
Q

What causes termination during translation.

A

-Proteins called release factors, which interact with STOP codons.
-RF1 recognises UAA and UAG
-RF2 recognises UAA and UGA
-RF3.GTP helps RF1 and RF2 carry out termination
-RRF and EF-G promote dissociation of the ribosome.

195
Q

Describe Catabolic pathways in metabolism.

A

Reactions which break down
-Degradative
-Energy producing
-Oxidative

196
Q

Describe Anabolic pathways in metabolism.

A

Reactions that build up.
-Biosynthetic
-Energy consuming
-Reductive

197
Q

How can +ve ΔG reactions be done?

A

-Increase temperature
-Increase the substrate concentrations
-Link an unfavourable +ve reaction with a favourable -ve reaction so that the reaction is overall -ve.

198
Q

What is the typical role of ATP?

A

Universal energy molecule

199
Q

What is the typical role of GTP?

A

Protein metabolism

200
Q

What is the typical role of CTP?

A

Lipid biosynthesis.

201
Q

What is the typical role of UTP?

A

Carbohydrate metabolism

202
Q

What is NAD?

A

Nicotinamide adenosine dinucleotide

203
Q

What is FAD?

A

Flavin adenosine dinucleotide

204
Q

What can be used to control metabolic pathways.

A

-Negative feedback
-Feed forward
-Isoenzymes
-Multienzyme complexes
-Compartmentalised
-Reciprocal regulation.

205
Q

Describe the Investment stage of glycolysis.

A

Glucose
↓ Hexokinase
Glucose-6-phosphate
↓
Fructose-6-phosphate
↓Phosphofructokinase
Fructose-1,6-bisphosphate

206
Q

Describe the splitting stage of glycolysis.

A

Fructose-1,6-bisphosphate is converted to either Glyceraldehyde-3-phosphate (by aldolase) or Dihydroxyacetone-phosphate, which is then converted to G3P by Triosephosphate isomerase.

207
Q

Describe the yield stage of glycolysis.

A

Glyceraldehyde-3-phosphate

↓ Glyceraldehyde-3-phsophate dehydrogenase

1,3-bisphosphoglycerate + NADH

↓Phosphoglycerate kinase

3-phosphoglycerate + ATP

↓Phosphoglycerate mutase

2-phosphoglycerate

↓Enolase

Phosphoenolpyruvate

↓Pyruvate Kinase

Pyruvate + ATP

208
Q

Why must metabolic pathways be tightly regulated?

A

-To link supply with demand
-To allow cells/organisms to responds to environmental changes
-To maintain a constant internal environment (homeostasis)
-To enable different tissues to interact (eg liver, adipose, muscle)

209
Q

How can glycolysis be controlled?

A

-Rate limiting enzymes
-Enzymes at the start or branch of the pathway
-Controlling the amount of an enzyme (slow)
-Controlling the activity of an enzyme through conformational change or phosphorylation (fast)

210
Q

How do high levels of AMP affect glycolysis?

A

Increase glycolysis.

211
Q

What suppresses/inhibits glycolysis?

A

-Low pH
-Citrate
-ATP
-Glucose-6-phosphate inhibits hexokimase

212
Q

What enzyme allows for pyruvate to enter the citric acid cycle, and how?

A

-Pyruvate dehydrogenase
-Which converts Pyruvate into Acetyl CoA and CO2, through oxidative decarboxylation

213
Q

What enzymes make up the pyruvate dehydrogenase complex?

A

-Pyruvate dehydrogenase (E1)
-Dihydrolipoyl transacetylase (E2)
-Dihydrolipoyl dehydrogenase (E3)

214
Q

What vitamins are required for the Citric acid cycle?

A

-B1 as thiamine pyrophosphate
-Riboflavin as FADH2
-Niacin as NAD

215
Q

What else can happen to pyruvate, other than entering the citric acid cycle?

A

-Converted to fatty acids (when ATP levels are high acetyl CoA diverted to a more efficient storage molecule)
-Converted to amino acids (During anabolic growth pyruvate is aminated to non-essential amino acids)
-Converted to lactate or ethanol (under anaerobic conditions to regenerate NAD+)

216
Q

What is the purpose of the citric acid cycle?

A

To harvest high-energy electrons from carbon fuels.

217
Q

Describe the citric acid cycle.

A

Oxaloacetate (4C) + S-CoA
↓Citrate synthase
Citrate (6C)
↓Aconitase
Isocitrate (6C)
↓Isocitrate dehydrogenase
⍺-Ketoglutarate (5C)
↓⍺Ketoglutarate dehydrogenase complex
Succinyl CoA (4C + S + CoA)
↓Succinyl CoA synthetase
Succinate (4C)
↓Syccinate dehydrogenase
Fumarate (4C)
↓Fumarase
Malate (4C)
↓Malate dehydrogenase
Oxaloacetate (4C)

218
Q

What can Citrate be converted to?

A

Fatty acids, sterols

219
Q

What can ⍺-Ketoglutarate be converted to?

A

Glutamate, which can then be converted to other amino acids or purines

220
Q

What can Succinyl CoA be converted to?

A

Porphyrins, heme, chlorophyll

221
Q

What can Oxaloacetate be converted to?

A

Aspartate, which can then be converted to other amino acids, purines, or pyrimidines

222
Q

What does the electron transport chain consist of?

A

-Complex I (NADH-Q oxidoreductase)
-Coenzyme Q (AKA Ubiquinone)
-Complex III (Q-cytochrome c oxidoreductase)
-Cytochrome C
-Complex IV (Cytochrome C oxidase)

223
Q

Describe Complex I (NADH-Q oxidoreductase) of the ETC.

A

-46 polypeptide chains
-Transfer of two high-potential electrons from NADH to FMN as it passes through its iron complex
-Electrons from FMNH2 are transferred to a series of FeS clusters
-Electrons from FeS clusters are shuttled to coenzyme Q

224
Q

Describe Coenzyme Q in the ETC.

A

-Q carries the electrons from NADH and FADH2
-Is hydrophobic and diffuses rapidly within the inner mitochondrial membrane.

225
Q

Describe Complex III (Q-cytochrome c oxidoreductase) in the ETC.

A

-Electrons transferred from QH2 to oxidised cytochrome c
-The mechanism that couples electron transfer from QH2 to cytochrome c is known as the Q cycle.

226
Q

Describe Cytochrome c in the ETC.

A

-Cytochrome c is present in all organisms with mitochondrial respiratory chains and has a highly conserved structure.
-Small soluble protein containing a c-type haem
-Carries one electron from Q-cytochrome c oxidoreductase to cytochrome c oxidase.

227
Q

Describe Complex IV (cytochrome c oxidase) in the ETC.

A

-4e- from cytochrome c are transferred to O2
-4H+ from matrix complete reduction of O2 to H2O
-4 more H+ are pumped across the membrane
-4H+ pumped from matrix to intermembrane space
-4 chemical protons removed from matrix.

228
Q

How does oxidative phosphorylation produce ATP?

A

The proton gradient produced by proton pumping during the electron transport chain is used to synthesize ATP. Protons flow down their concentration gradient into the matrix through the membrane protein ATP synthase, causing it to spin (like a water wheel) and catalyze conversion of ADP to ATP.

229
Q

How does Cyanide affect oxidative phosphorylation?

A

Binds to Complex IV and blocks oxidative phosphorylation

230
Q

How does salicylate affect oxidative phosphorylation?

A

-Forms pores in the inner membrane, meaning protons pass through without generating ATP.

231
Q

What range should blood glucose be?

A

4-6mmol/L in the plasma

232
Q

When does hypoglycaemia occur?

A

-During starvation
-Insulin overdose
-During and after exercise

233
Q

How much metabolised glucose does the brain use daily?

A

75% (~120g/day)

234
Q

When does hyperglycaemia occur?

A

-Post prandial
-Inadequate insulin administration

235
Q

What are the sources of gluconeogenesis?

A

-Pyruvate/lactate
-Glycerol
-Citric acid cycle intermediaries
-Amino acids

236
Q

When is gluconeogenesis activated?

A

-Initiated 2 hours post prandially
-Preserves glycogen for emergencies
-Maximum 8-12 hours
-At high intensity exercise

237
Q

Where does gluconeogenesis occur?

A

Principally in the liver.

238
Q

Describe the Cori cycle of gluconeogenesis.

A

Lactate in the muscle is transferred through the blood to the liver, where it is converted to pyruvate, which can then either be converted to glucose or CO2 and H2O

239
Q

Describe the Alanine cycle of gluconeogenesis.

A

Lactate in the muscle is converted to pyruvate, which is then aminated to Alanine. This alanine then travels through the blood to the liver where it is converted to pyruvate where it can then be converted to either glucose or CO2 and H2O

240
Q

What are the three energetically unfavourable reactions that occur to convert pyruvate to glucose?

A

-Pyruvate to PEP
-F1,6bisphosphate to F6Phosphate
-G6P to glucose

241
Q

Describe the reaction that occurs during gluconeogenesis when pyruvate is converted to PEP.

A

-Pyruvate is converted to oxaloacetate utilising ATP, CO2, through the enzyme pyruvate carboxylase (using coenzyme Biotin)
-Oxaloacetate is converted to Phosphoenolpyruvate (PEP) utilising GTP and producing CO2, through the enzyme PEP carboxykinase

242
Q

Describe the reaction that occurs during gluconeogenesis when Fructose1,6bisphosphate is converted to Fructose6phosphate.

A

F1,6bisphosphate is converted to F6Phosphate using the enzyme Fructose-1,6-bisphosphatase, using water and releasing a phosphate group.

243
Q

Describe the reaction that occurs during gluconeogenesis when Glucose-6-phosphate is converted to glucose.

A

G6P is converted to glucose using the enzyme Glucose-6-phosphatase, requiring H2O and releasing a Phosphate group

244
Q

Gluconeogenesis requires ATP and GTP, but where does this come from?

A

-Lipid oxidation
-Amino acid catabolism

245
Q

Why do AMP and ADP inhibit gluconeogensis?

A

-Indicates a low energy status
-need to send pyruvate into TCA cycle to get energy

246
Q

Why do citrate and acetyl CoA activate gluconeogenesis?

A

-Indicate high energy status and plenty of biosynthetic precursors
-Favours conversion of pyruvate to G6P for glycogen synthesis.

247
Q

Describe Muscle Glycogen

A

~2% of muscle weight is glycogen
-Largest glycogen store in body
-Glucose released as glucose-6-phosphate just for muscle

248
Q

Describe Liver Glycogen

A

~10% by weight is glycogen
-Controls blood glucose levels
-Glucose is released for other cells

249
Q

How much glycogen is stored in the body at any given time?

A

On average 450g, which is ~18 hours of supply

250
Q

Describe Glycogenesis.

A

Glucose-6-phosphate
↓Phosphoglucomutase
Glucose-1-Phosphate
↓ UDP-glucose pyrophosphorylase + UTP
UDP-Glucose
↓ Glycogen synthase
Glycogen (N+1)

251
Q

What enzyme is responsible for glyogen branching?

A

Amylo-(1,4 -> 1,6)-transglycosylase

252
Q

What does glycogen synthase require?

A

Glycogenin, which limits the size of glycogen, as well as acting as a primer for glycogen synthase.

253
Q

What enzyme erodes the end chains of glycogen in glycogenolysis?

A

Glycogen phosphorylase, which utilises the energy of the glycosidic link to phosphorylase (using inorganic P).

254
Q

Describe the stages of glycogenolysis.

A

1 - Erosion of end chains by glycogen phosphorylase, utilising the energy of the glycosidic bond. Glycogen phosphorylase stops at the 4th glucose in a chain
2 - ⍺(1->4)transglycosylase removes 3 glucose off a branch and adds it to main chain, and ⍺(1->6)transglycosylase removes final off the branch.
3 - Phosphoglucomutase converts glucose-1-phosphate to glucose-6-phosphate which can then be used in glycolysis.

255
Q

Which enzyme triggers glycogenesis?

A

Insulin

256
Q

Which enzyme triggers glycogenolysis?

A

Glucagon

257
Q

Give some symptoms of Glycogen storage diseases (GSD)

A

-Muscle cramp
-Muscle weakness
-General tiredness
-Hypoglycaemia
-Hepatomegally

258
Q

What causes Type 0 Glycogen storage disease?

A

-Liver glycogen synthase deficiency
-Meaning that glucose-1-phosphate is not converted to glycogen, leading to hyperglycaemia and hypoglycamia

259
Q

What causes Type I Glycogen storage disease (Von Gurke’s)?

A

-Glucose-6-phosphatase deficiency
-Meaning that Glucose-6-phosphate is not converted back to glucose, leading to hypoglycaemia and hepatomegally.

260
Q

What causes Type III Glycogen storage disease (Cori’s)?

A

-Amylo-1,6-glucosidase deficiency
-Second active site of debrancher is affected, meaning that glycolysis is blocked at the branches, leading to hepatomegally and hypoglycaemia.

261
Q

What causes Type IV Glycogen storage disease (Andersen’s)?

A

-Amylo-(1,4->1,6)-glucosidase deficiency
-Lack of this brancher enzyme leads to long chain glycogen, which makes it water insoluble and leads to slow glucose release.
-Leads to hepatomegally and hypoglycaemia.

262
Q

What causes Type V Glycogen storage disease (McArdle’s)

A

-Muscle phosphorylase deficiency
-Unable to release terminal sugars, meaning that muscle requirements not met during exercise, leading to muscle cramps and weakness.

263
Q

During exercise, what happens to glucose uptake, glycogen breakdown and synthesis in the muscle?

A

Glucose uptake↑
Glycogen breakdown↑
Glycogen synthase↓

264
Q

At rest, what happens to glucose uptake, glycogen breakdown and synthesis in the muscle?

A

Glucose uptake↓
Glycogen breakdown↓
Glycogen synthesis↑

265
Q

After a meal, what happens to glucose uptake, glycogen breakdown and synthesis in the liver?

A

Glucose release↓
Glycogen breakdown↓
Glycogen synthesis↑

266
Q

During fasting/exercise, what happens to glucose uptake, glycogen breakdown and synthesis in the liver?

A

Glucose release↑
Glycogen breakdown↑
Glycogen synthesis↓

267
Q

What 3 hormones regulate glycogen metabolism?

A

-Epinephrine (provides energy in an emergency)
-Glucagon and Insulin (fine control of blood sugar levels)

268
Q

How does Epinephrine affect its target cells?

A

-Epinephrine acts via Gprotein linked receptor to initate a cAMP-dependent phosphorylation cascade, leading to stimulation of glycogen breakdown (and inhibits glycogen synthesis).

269
Q

How does Insuilin affect its target cells?

A

Insulin binds to a receptor tyrosine kinase, which phosphorylates an Insulin response substrate, leadung to a cascade that phosphorylates glycogen synthase. This activates it leading to more glycogen synthesis

270
Q

What does the GLUT2 transporter control?

A

GLUT2 mediates secretion of insulin by pancreatic β-cells when blood sugar is high.

271
Q

Describe Type 1 diabetes

A

~10% of diabetes cases in UK
-Absolute insulin deficiency due to βcell destruction, usually autoimmune response.
-Treated with exogenous insulin injection or pancrease/islet transplant

272
Q

Describe Type 2 diabetes

A

~90% diabetes cases in UK
-Stage 1 is hyperinulinemia/insulin resistance
-Stage 2 is beta cell “hideaway” to avoid death
-Reversible by diet and exercise
-Abnormal phenotypes in pancreas of Type 2

273
Q

What can hyperglycaemia lead to?

A

↑Glycogen breakdown
↑Fat breakdown (generating ketone bodies)
↑Gluconeogenesis
↑Protein breakdown

274
Q

What are fats stored as, and mainly in what tissue?

A

Stored mainly as triglycerides, mainly in white adipose tissue

275
Q

What is a polyunsaturated fat?

A

A fatty acid chain with multiple double bonds.

276
Q

What are the 3 stages of fat catabolism?

A

1 Lipolysis
2 Activation
3 Oxidation

277
Q

What enzymes are involved in Lipolysis?

A

-Adipose triglyceride lipase (ATGL)
-Hormone-sensitive lipase (HSL)
-Monoglyceride lipase (MGL)

278
Q

What is glycerol converted to following lipolysis?

A

D-Glyceraldehyde-3-phosphate, which can then can be converted to glucose or pyruvate.

279
Q

Describe the steps of conversion from glycerol to D-Glyceraldehyde-3-phosphate

A

Glycerol
↓ ATP
L-Glycerol-3-phosphate
↓NAD+
Dihydroxyacetone phosphate
↓
D-Glyceraldehyde-3-phosphate

280
Q

Describe Fatty acid activation

A

Fatty acid is combined with CoA by Acyl CoA synthetase, utilising the energy of pyrophosphate hydrolysis

281
Q

What is the acyl group transported into the mitochondria as?

A

Acyl-Carnitine

282
Q

What process derives energy from acyl groups?

A

Beta-oxidation

283
Q

Describe the steps of beta-oxidation of an acyl group.

A

1 - Activated fatty acid is oxidised by FAD (and acyl-CoA dehydrogenase)
2 - Hydration by enoyl CoA hydratase
3 - Dehydrogenate to oxidise the beta carbon using NAD and L-3-hydroxyacyl CoA dehydrogenase
4 - Cleavage/thiolysis by 3-ketoacyl thiolase
5 - Repeat until there are only 2 carbons, to maximise amount of Acetyl CoA produced.

284
Q

What is the ATP yield from complete oxidation of glucose vs palmitate

A

32 ATP (0.18ATP per dalton) vs 108 ATP (0.42ATP per dalton)

285
Q

Describe Ketone formation, and why is it significant for survival?

A

-Produced mainly in liver
-Can be used as energy source by brain and heart in starvation

286
Q

How does Malonyl CoA affect fatty acid oxidation?

A

It inhibits the process

287
Q

Where does fatty acid synthesis occur in the cell?

A

In the cytoplasm

288
Q

Where does fatty acid degradation occur in the cell?

A

Mostly in the mitochondrial matrix

289
Q

What are essential amino acids?

A

Amino acids that cannot be synthesised by humans thus supplied only from diet.

290
Q

What are non-essential amino acids?

A

Amino acids that can be synthesised by humans if not supplied in diet.

291
Q

Which amino acids are the essential amino acids?

A

Isoleucine, Leucine, Valine, Phenylalinine, Tryptophan, Histidine, Methionine, Lysine, Tyrosine

292
Q

Where does protein catabolism occur?

A

Mainly in the liver.

293
Q

What do transaminases do?

A

Convert amino acids into keto-acids.

294
Q

What reaction does aspartate aminotransferase catalyse?

A

The conversion of Oxaloacetate and Glutamate into Aspartate and ⍺-ketoglutarate

295
Q

What amino acids can pyruvate be converted into?

A

Alanine, Valine and Leucine

296
Q

What amino acids can oxaloacetate be converted into?

A

Aspartate, Asparagine, Methionine, Threonine, Isoleucine, Lysine

297
Q

What amino acids can ⍺-ketoglutarate be converted into?

A

Glutamate, Glutamine, Proline, Arginine

298
Q

What amino acids can Phosphoenolpyruvate and Erythrose 4-Phosphate be converted into?

A

Phenylalanine, Tyrosine, Tryptophan

299
Q

What amino acid can Ribose 5-phosphate be converted into?

A

Histidine

300
Q

What amino acids can 3-Phosphoglycerate be converted into?

A

Serine, Cysteine, Glycine

301
Q

What metabolic intermediates can amino acids be converted to?

A

-Acetyl CoA
-Acetoaceetyl CoA
-Pyruvate
-⍺-keto glutarate
-Succinyl CoA
-Fumarate
-Oxaloacetate

302
Q

Why do we like ethanol?

A

Increases response to the GABA neurotransmitter

303
Q

At what rate is ethanol metabolised in the liver?

A

100mg/kg/hr

304
Q
A
305
Q

At what ethanol concentration does death occur?

A

500mg/100ml

306
Q

Describe the steps of ethanol metabolism.

A

Ethanol
↓Alcohol dehydrogenase + NAD
Acetaldehyde
↓ Alcohol dehydrogenase + NAD + H2O
Acetic acid

307
Q

What negative effects does ethanol consumption have on the body?

A

-Depletes NAD+
-Excessive NADH inhibits gluconeogenesis and Citric Acid Cycle.
-Symptoms include Acidosis, Ketosis, Hypoglycaemia, Lipogenesis

308
Q

At what stage of fatty liver disease is liver transplant needed or death occurs?

A

Alcohol-associated Cirrhosis (AC) and Hepatocellular Carcinoma (HCC)

309
Q

In gene expression in E.coli, which termination sequences for translation are recognised by RF1?

A

UAA and UAG

310
Q

what are the main enzymes used in protein catabolism?

A

Transaminases

311
Q

In E.coli, what ion is RNA polymerase dependent on in order to begin transcription?

A

Mg2+

312
Q

What must insulin do in order to be stored in high concentrations in the pancreas?

A

It must form a hexamer

313
Q

Describe the steps of the investment stage of glycolysis.

A

1 - Phosphorylating glucose (Using ATP)
2 - Converts the aldopyranase to a ketofuranose
3 - Further phosphorylates the ketofuranose

314
Q

Describe the steps of the splitting stage o

A

1 - Splitting the single 6-carbon molecule into two 3-carbon molecule
2 - One of the split molecules is not G3P, so must be converted to this by triosephosphate isomerase

315
Q

Describe the steps of the yield stage of yield stage of glycolysis.

A

1 - Glyceraldehyde-3-Phosphate is dehydrogenated and a free phosphate is gained
2 - Phosphoglycerate kinase is used to harvest a phosphate, producing ATP
3 - Phosphate is transferred to central carbon by phosphoglycerate mutase
4 - Energy potential is increased by removing water (using enolase)
5 - Pyruvate kinase is used to convert PEP to pyruvate and ATP

316
Q

Give the products formed during the citric acid cycle, starting with Citrate.

A

1 Citrate
2 Isocitrate
3 ⍺-ketoglutarate
4 Succinyl CoA
5 Succinate
6 Fumarate
7 Malate
8 Oxaloacetate

317
Q

What enzyme converts Citrate to Isocitrate?

A

Aconitase

318
Q

What enzyme converts Isocitrate to ⍺-ketoglutarate?

A

Isocitrate dehydrogenase (with NAD+)

319
Q

What enzyme converts ⍺-ketoglutarate to Succinyl CoA

A

⍺-ketoglutarate dehydrogenase complex (with NAD+ and CoA)

320
Q

What enzyme converts Succinyl CoA to Succinate?

A

Succinyl CoA synthetase (using GDP + Pi)

321
Q

What enzyme converts Succinate to Fumarate?

A

Succinate dehydrogenase

322
Q

What enzyme converts Fumarate to Malate?

A

Fumarase (with H2O)

323
Q

What enzyme converts Malate to Oxaloacetate?

A

Malate dehydrogenase (with NAD+)

324
Q

What enzyme converts Oxaloacetate to Citrate?

A

Citrate synthase

325
Q

In SDS-PAGE, the migration of the protein is …

(MW = the molecular weight of the protein in Daltons)

A

proportional to log(MW)

326
Q

If a reaction is anaplerotic, what does the reaction do?

A

Create intermediates for metabolism reactions.

327
Q

An non-active enzyme without its coenzyme and will be activated once combined with its coenzyme. This molecule is known as __________?

A

Apoenzyme

Year 1 Semester 1 (4 decks)

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