Biochem Midterm I - Oct 11th Flashcards
Describe Proteins
- Covalently linked amino acids in an approx 20 amino acid chain
- AKA polypeptides
- Amino acids connected by peptide bonds between C-N, an Amino-terminal end (NH3) and a carboxyl end (O=C-O)
Describe carbohydrates
- Structure and Composition: Carbohydrates are organic molecules made up of carbon, hydrogen, and oxygen, typically with a hydrogen-to-oxygen ratio of 2:1, resembling that of water (H₂O).
- Types: They are classified into three main categories: monosaccharides (simple sugars), disaccharides (two sugar units), and polysaccharides (long chains of sugar units), each serving different roles in biological processes.
- Functions: Carbohydrates provide energy for living organisms, serve as structural components in cells (like cellulose in plants), and are involved in cell recognition and signalling.
Describe polysaccharides
large, complex carbohydrates composed of long chains of monosaccharide (single sugar) units linked together by glycosidic bonds. These bonds form through dehydration synthesis, resulting in various structures such as starch, glycogen, and cellulose, each serving different functions in living organisms. Their unique configurations contribute to properties like energy storage, structural support, and biological recognition.
Describe esters in the context of lipids
The chemical compounds formed when fatty acids react with alcohol, typically glycerol, through a condensation reaction. This reaction results in the formation of triglycerides, which are the main constituents of body fat in humans and other animals, as well as vegetable fat.
What are the three major types of lipid esters?
- Glycerophospholipid: glycerol ester
- Sphingolipids: sphingosine ester
- Steroids: Derivative of waxes making nonpolar esters w a long chain
Describe the structure of P-glycoprotein
- ATP-binding cassette (ABC)
- Contain two homologous halves, each containing transmembrane segments (x6 per, aka MSD1 and MSD2) and a nucleotide-binding domain (NBD)
- The MSDs form a hydrophobic channel that facilitates the efflux of substrates
- The NBDs interact with ATP to cause conformational changes in the R complex, enabling the transport cycle that effectively expels substrates from the intracellular milieu, thus playing a crucial role in multidrug resistance.
Regarding the two members of the ATP-binding cassette (ABC) superfamily:
____ functions as an efflux pump, expelling drugs and toxins, while ____ acts as a chloride channel regulating ion and water transport in epithelial tissues. Both utilise ATP hydrolysis to drive conformational changes, but ____includes unique regulatory elements; and its dysfunction causes cystic fibrosis, whereas altered ____ function can lead to multidrug resistance in cancer.
P-gp
CFTR
CFTR
P-gp
What are the three types of systems?
- Open: Matter and energy are freely exchanged with the surroundings in an open system
- Closed: Only energy is exchanged with the surroundings in a closed system
- Isolated: Neither matter nor energy is exchanged with the surroundings in an isolated
Describe the first law of thermodynamics
- Energy (E) can neither be created nor destroyed
- Energy can only be converted from one form to another
- q = heat absorbed from surrounding, w = work done to surrounding
- ∆E = EFinal - EInitial = q * w
- Energy has an effect on enthalpy (heat content, H)
- H = E + PV
- If constant pressure:
- ∆H = ∆E + P∆V = q - w + P∆V = qv for the volume change is insignificant in most biological systems
What is the second law of thermodynamics?
- All spontaneous processes in the universe tend toward disorder (entropy, S) in the absence of energy input
- Entropy definition: The degree of randomness of a system
- S = KBlnW
- Where W is the number of energetically equivalent ways and KB is the Boltzmann constant (1.38x10-23 j/K)
Compare exothermic and endothermic enthalpy
- Exothermic
- Reaction releases heat
- Has a -∆H
- Example: wearing a winter coat to keep your body warm during cold weather - Endothermic
- Reaction absorbs heat
- Has a +∆H
- Example: Absorbing heat into your skin from a heated blanket
Describe Gibbs free energy
- The difference between enthalpy (H) and entropy (S) of a system at a given temperature
- G = H - TS
- At equilibrium, ∆G = 0, the rate of formation of products and reactants is equal
Exergonic:
- ∆G < 0
- Seen in forward rxns
- Reaction is favourable and spontaneous
Endergonic:
- ∆G > 0
- Seen in reverse rxns
- Rxn is unfavourable and non-spontaneous
Describe the difference between Gibbs free energy (∆G) and Standard gibbs free energy (∆Gº)
In essence, ΔG is context-specific, while ΔGº provides a standard benchmark for evaluating reactions. Other conditions include:
- pH = 7
- [H2O] = 55.5 M
- Mg2+ = 1 mM
Describe Standard gibbs free energy (∆Gº)
ΔGº (Standard Gibbs Free Energy Change) represents the free energy change under standard conditions (1 M concentrations, 1 atm pressure, and typically 25°C), serving as a reference for comparing reaction favorability.
Describe the relationship between ∆Gº and Keq
Describe coupled reactions
- If a rxn is endergonic, it can be coupled to an exergonic rxn to become overall favourable
- ATP hydrolysis is a common coupling rxn as its ∆Gº = - 30.5 kJ/mol
- When coupling a rxn, you must add up the free energies of the separate rxns to find the overall free energy
Describe water molecules
- Water is a polar molecule
- The six outer orbital electrons of oxygen atom are distributed in the four sp3 hybrid orbitals
- The two hydrogen atoms are in the two corners of the orbital tetrahedron
- The angle between the two O-H bonds is 104.3º instead of the usual 109.5º because the lone pairs of electrons on water exert greater repulsive forces than the bonded hydrogens.
Describe H bonding
- Hydrogen bonding occurs when an H atom is covalently bonded to an e-neg atom and in close proximity to another e-neg atom (ex. O or N)
- Hydrogen is “shared” between two e-neg atoms
____ is the substance that dissolves a solute, typically present in a greater amount, and is the medium in which the solution occurs (e.g., water in saltwater). _____ is the substance that is dissolved in the solvent, usually present in a smaller amount (e.g., salt in saltwater). Together, they form a solution
Solvent
Solute
Each ion is surrounded by one or more concentric shells of oriented solvent molecules, which essentially spread the ionic charge over a much larger volume. Such ions are said to be ____ or, when water is the solvent, to be _____
solvated
hydrated
Describe hydrophobic interactions
- Non-polar substances tend to aggregate in water. It results from the strong tendency of water to exclude nonpolar groups or molecules
- The introduction of hydrophobic substance causes increased water surfaces and disrupts original water bonds
- The essence of hydrophobic effect is to minimise the cost of disrupting the water hydrogen bonds and increase H2O entropy
- For aggregation process: ∆G<0
Describe the aggregation of hydrophobic substances in water
Water excludes non-polar molecules to avoid disrupting its stable hydrogen bonds. To reduce this disruption, hydrophobic substances aggregate, minimising their exposed surface area to water. This aggregation increases water’s entropy, as fewer ordered water “cages” are needed around the clustered hydrophobic molecules, allowing more water molecules to move freely.
Describe amphiphilicity
- Molecules that are attracted to both polar and nonpolar environments
- All amphiphiles form micelles or bilayers
- The formation of micelles or bilayers are _______ ______
- The size of a micelle and the number of surfactant molecules it contains are influenced by the size of the ____ ____ group and the length of the ____ ____.
thermodynamically favourable
polar head
nonpolar tail
A larger polar head group tends to result in _____ micelles due to increased steric hindrance, while longer nonpolar tails promote ____ micelles by enhancing hydrophobic interactions. This balance between head group size and tail length determines the optimal micelle formation, impacting applications like drug delivery and emulsification
smaller
larger
Describe detergents
- A special group of amphiphile: the molecule which has both polar and nonpolar segments
- Structurally the simplest version of lipids
- A detergent forms micelles in water only when its conc. is above the critical micellar concentration (CMC)
- Depending on the head group, the detergents can be divided into ionic and non-ionic.
- Ionic detergents are more powerful at solubilising proteins, or for washing your clothes
Describe amphipathicity
Molecules that contain both polar and nonpolar groups (i.e fatty acids)
How are amphilicity and amphipathicity different
Amphiphilic refers to a molecule that has both hydrophilic (water-attracting) and hydrophobic (water-repelling) parts and has an affinity for both polar and non-polar environments. It’s typically used in the context of how molecules interact with water and oils, such as surfactants.
Amphipathic also refers to molecules with both hydrophilic and hydrophobic regions, but the term emphasizes the structural division of these regions within the molecule. It’s often used in a biological context, such as describing lipids in membranes.
What are the acid a propos equations?
pH = -log[H+]
pKw = pH + pOH = 14
HA ⇌ H+ + A-
Describe [H+]
This denotes the concentration of hydrogen ions (protons) in a solution. It is a measure of acidity, and higher [H+] concentrations correspond to lower pH values.
Describe [HA]
This represents the concentration of the undissociated weak acid, where “HA” signifies the acid itself (e.g., acetic acid). It is the form of the acid that is not dissociated into ions in solution.
Describe [A-]
This indicates the concentration of the conjugate base of the weak acid, formed when the acid donates a proton. For example, if HA is acetic acid, then [A−] would refer to the acetate ion (CH3COO-).
The pH at the midpoint of a titration provides the ___ value
pKa
Define a weak acid
An acid that IS NOT completely dissociated in the water solt.
Describe buffers
The solt. which contain at least one pair of weak acid and its conjugate base
- The best buffering capacity occurs when there is [HA] = [A-], in other words when pH= pKa
- pKa changes when solt. temp changes
Describe the dissociation constant of a weak acid
Ka = [H+][A-]/[HA]
- A higher Ka value indicates a stronger acid (greater dissociation), while a lower Ka value indicates a weaker acid
- Usually it is products on top and reactants on the bottom
Describe the titration of phosphatic acids
The titration of phosphoric acid (H3PO4), a triprotic acid, involves sequentially adding a strong base, such as sodium hydroxide (NaOH), to determine the pKa values corresponding to its three dissociation steps. Each dissociation yields a conjugate base: H2PO4−, H2PO42- and PO43−, with distinct equivalence points observable in the titration curve. The resulting pH changes at each equivalence point are indicative of the buffer regions, reflecting the acid-base equilibrium of the phosphate species. The titration curve typically displays a characteristic sigmoidal shape, allowing for the precise determination of pKa values and the acid’s overall strength.
Describe the significance of the x-axis of a titration graph
- Equivalence Points: The inflection points on the sigmoidal curve indicate the volumes of titrant where specific protons from phosphoric acid have been completely neutralised.
- Buffer Regions: Between equivalence points, the curve shows gradual pH changes, reflecting the buffering capacity of the acid-base pairs present.
- Titration Progress: The x-axis illustrates the relationship between titrant volume and pH changes, helping to assess the strength and behaviour of the acid throughout the titration.
Describe zwitterions
Dipolar (containing both + and - charges but overall neutral) amino acids. The predominant form of amino acids in pH neutral conditions. The + group is NH3+ and the - group is COO-. This dual charge allows zwitterions to exhibit unique properties, such as high solubility in water and the ability to act as both acids and bases, influencing their behaviour in biological systems and chemical reactions.
Which amino acids are charged?
Asparagus Glue Lyses Argentinian Hispanics
Asp
Glu
Lys
Arg
His
Which amino acids are hydrophilic?
Ser
Thr
Cys
Asn
Gln
Seriously Thrown Cysts Assassinate Gringos
Which amino acids are hydrophobic
Gly
Ala
Pro
Val
Leu
Ile
Met
Gliding Allow Professionally
Valued Losers Illegitimate Methods
Which amino acids are aromatic?
Phe
Tyr
Trp
Phoenix’s Try Trips
Which amino acids are non-polar?
Gly
Ala
Pro
Val
Leu
Ile
Met
Phe
Tyr
Trp
Gliding Allow Professionally
Valued Losers Illegitimate Methods for Phoenix’s Trying Trips
Which amino acids have uncharged polar side chains?
Ser
Thr
Asn
Gln
Some Asinine Threesomes Golong
Which amino acids contain ionised polar side chains?
Lys
Arg
Asp
Glu
Asparagus Glue Lyses Argentinians
What amino acids contain partially ionised polar side chains?
His
Tyr
Cyst
His Tyred Cyst
Describe the formation of cystine
Describe proline
Why does the ring structure of proline restrict protein geometry?
The cyclic nature of proline results in a rigid structure that restricts the flexibility of the polypeptide chain. Unlike other amino acids, proline’s side chain is part of the backbone, limiting the possible torsion angles around the peptide bond
- All standard amino acids are chiral except for ____
- All L-amino acids are also S-amino acids except for ____
- _____ has four stereoisomers since it contains two different chiral centres
glycine
cystine
Isoleucine
What bioactive molecules are the derivatives of the following amino acids, respectively?:
1. Glu
2. His
3. Tyr
- GABA
- Histamine
- Dopamine or Thyroxine
Describe the dimerisation of glutathione
Two molecules of the tripeptide glutathione (composed of glutamate, cysteine, and glycine) combine to form glutathione disulfide (GSSG) through the oxidation of cysteine’s thiol groups, creating a covalent disulfide bond. This reversible dimerization is key to cellular redox balance. Reduced glutathione (GSH) functions as an antioxidant, while its oxidised form (GSSG) can be converted back to GSH by glutathione reductase, maintaining redox homeostasis during oxidative stress.
The OH groups of Ser, Thr, Tyr are often _____
The NH2 group of Lys is often _____ (H → CH3-C=O)
The ______ is a major amino acid in collagen
The modification of a side chain is called a ______ event
phosphorylated
acetylated
hydroxyproline
posttranslational
Describe how the property of a polypeptide is determined by side chains of amino acids
- By formation of peptide bonds, the ionisable states of -NH2 and COOH are lost and only one -NH2 and one COOH group are left at the end (side chain may also contain the ionisable groups)
- The alpha carbon and the peptide bonds form the backbone of a polypeptide
Describe Hydrophobic side chains
(like those of leucine, isoleucine, and phenylalanine) tend to cluster together in the interior of proteins, helping to stabilise the protein’s three-dimensional structure.
Describe Hydrophilic side chains
(like those of serine, threonine, and glutamine) often participate in hydrogen bonding and are usually found on the protein surface, interacting with water or other polar molecules.
Describe Charged side chains
(like lysine, arginine, and glutamate) play critical roles in forming salt bridges, participating in enzyme catalysis, or interacting with the surrounding environment, depending on their charge at physiological pH.
Describe the Isoelectric point
The isoelectric point (pI) is the pH at which a molecule, such as an amino acid or protein, carries no net electrical charge. At this pH, the concentrations of positively and negatively charged forms of the molecule are equal, resulting in a state of electrical neutrality.
Describe salting out
- A technique for precipitating proteins by increasing salt concentration, usually with ammonium sulphate. As salt concentration rises, water molecules are drawn to the salt ions, reducing the protein’s hydration shell and solubility, causing aggregation and precipitation.
- Used in the initial steps of protein purification to remove unwanted proteins and concentrate the target protein.
Describe Ion exchange chromatography
- Separates proteins based on their net charge at a specific pH.
- In anion exchange, negatively charged proteins bind to a positively charged matrix, while in cation exchange, positively charged proteins bind to a negatively charged matrix.
- Proteins are eluted by increasing salt concentration or adjusting pH to disrupt electrostatic interactions. - It is commonly used to purify proteins with known charge properties, such as separating proteins with slightly different isoelectric points (pI).
Describe Size-Exclusion Chromatography (Gel Filtration)
- Separates proteins based on size using a stationary phase of porous beads. Larger molecules elute first because they cannot enter the pores, while smaller molecules elute later as they travel through the pores.
- The beads contain specific-sized pores, allowing this separation.
- This technique is commonly used for desalting, buffer exchange, and separating proteins with significant differences in molecular weight, as well as in the final purification steps to remove aggregates or contaminants of varying sizes.
How do you calculate the sedimentation coefficient
What are the five steps of protein sequencing?
Describe Tandem Mass Spectrometry (MS/MS)
An advanced analytical technique used to identify and characterise biomolecules, particularly proteins and peptides, based on their mass-to-charge ratio (m/z). Here’s how it works:
1. Ionisation
* This process converts molecules in the sample into charged ions, which can be manipulated in the mass spectrometer.
2. First Mass Analysis (MS1)
* The generated ions are then introduced into the first mass spectrometer (MS1), where they are separated based on their m/z ratios.
3. Fragmentation
* Selected precursor ions are then fragmented in a collision cell, where they collide with neutral gas molecules. This collision causes the ions to break apart into smaller fragments, which provides structural information about the original molecules.
4. Second Mass Analysis (MS2)
* The resulting fragment ions are then analysed in a second mass spectrometer (MS2). This stage measures the m/z ratios of the fragment ions, allowing for the determination of their structure and the reconstruction of the original molecule’s sequence.
What is the difference between a type I and type II beta turn?
- Type I beta turn, the carbonyl oxygen of the second residue points inward, while in a type II beta turn, the carbonyl oxygen points outward. Additionally, type II turns often feature glycine as the third residue to avoid steric clashes.
- Only Type I β-turns are stabilized by hydrogen bonds, not Type II β-turns.
Describe the PII helix
A left-handed helical structure, prominent in proteins like collagen, where it forms a triple helix. The PII helix is stabilised by specific hydrogen bonds, and collagen’s composition—rich in glycine, proline, and hydroxyproline—ensures the formation of this unique structure.
Describe Cyclic Symmetry:
Cyclic symmetry refers to a circular arrangement of protomers around a central axis, allowing for rotational symmetry. For example, a protein with a 6-fold cyclic symmetry has six identical subunits arranged in a ring, contributing to the formation of structures like viral capsids or symmetric enzymes.
Describe Dihedral symmetry
- Combines both rotational and mirror symmetry. It describes arrangements where protomers are organized in such a way that there are symmetrical relationships between pairs of protomers, often resulting in more complex quaternary structures.
Describe Tetrahedral Symmetry:
Tetrahedral symmetry describes a three-dimensional arrangement where protomers are positioned at the corners of a tetrahedron. This symmetry is characteristic of certain viral capsids and multi-subunit proteins, facilitating a stable structure that can withstand environmental stresses
Describe zinc finger motifs
- A type of domain motif functional bending
- A beta sheet’s cystines bind with the histidines of an alpha helix via a zinc bridge to form a finger looking bend between the two
- This domain motif then binds to the major groove of DNA and helps regulate transcription
Define hydropathy
- The study of the hydrophobic and hydrophilic properties of molecules
- Quantified using a hydropathy index, which measures the relative hydrophobicity or hydrophilicity of amino acids; positive values indicate hydrophobicity, while negative values indicate hydrophilicity
- Helps predict membrane-spanning regions of proteins
What are the steps of protein de- and re-naturation
Describe Hsp70 in the context of E. coli
Hsp70 proteins, such as DnaK in Escherichia coli, play a crucial role in the folding of nascent polypeptides during their synthesis on ribosomes. These chaperones specifically recognise exposed, hydrophobic regions of polypeptides, preventing nonproductive associations and ensuring the polypeptide remains unfolded until productive folding interactions can occur. Hsp70 consists of two main domains: a 44-kDa N-terminal ATP-binding domain that regulates its activity and an 18-kDa central domain that binds to hydrophobic residues. By stabilising nascent chains, Hsp70 facilitates correct folding processes critical for protein functionality, especially under stress conditions that can induce misfolding.
What are the four steps in the functional cycle of GroEL/ES
Describe Ubiquitin-dependent protein degradation
Describe protein triage
- Definition: Cellular process for assessing and managing newly synthesised proteins to ensure proper folding and functionality.
- Folding on Ribosomes: Proteins begin to fold on ribosomes; some may misfold or remain partially folded.
- Role of Molecular Chaperones: Chaperones like Hsp70 and chaperonins (e.g., GroEL/GroES) bind to nascent or misfolded proteins, preventing aggregation and assisting in proper folding.
- Quality Control Mechanisms: Detect misfolded proteins, which may be targeted for degradation via the ubiquitin-proteasome system or redirected for refolding attempts.
- Outcome of Protein Triage: Determines whether proteins achieve native conformations, are refolded, or are degraded, ensuring cellular proteostasis and function, especially under stress conditions
Describe how protein disulphide isomerase catalyses disulfide interchange
