Chapters 1-3 Flashcards
1
Q
What are the ways in which biochemical reactions differ from chemical reactions?
A
Aqueous Faster Require energy (ATP) Localized in the cell Part of multi-step pathway Regulated
2
Q
What are the four types of biomolecules?
A
Lipids
Proteins
Carbohydrates
Nucleic Acids
3
Q
What is the monomer for lipids?
A
Fatty acids
4
Q
What is the monomer for proteins? What is the bond called?
A
Amino acids
Peptide bond
5
Q
What is the bond for carbohydrates?
A
Glycosidic bonds
6
Q
What is the difference between a nucleoside and a nucleotide?
A
Nucleotide contains a Phosphate
7
Q
What is the bond for nucleic acids?
A
Phosphodiester bonds
8
Q
What are the reactions called when monomers are made into polymers and vice versa?
A
Condensation- monomers are made into polymers and release a small molecule (water if dehydration reaction)
Hydrolysis- water ruptures the chemical bond
9
Q
What are the two categories of DNA bases and which bases are in which category?
A
Purines
Adenine & Guanine
Pyrimidines
Thymine & Cytosine
10
Q
What are the complementary base pairs in DNA and how many hydrogen bonds can each make?
A
Guanine and Cytosine → 3 hydrogen bonds
Adenine and Thymine → 2 hydrogen bonds
11
Q
What are the three differences between RNA and DNA?
A
Single-stranded
Ribose (vs deoxyribose)
Uracil (replace CH3 with H)
12
Q
Describe the three main molecular models
A
Space-filling - Realistic
Ball-and-stick - Bonding arrangements
Skeletal - Simple, good for large molecules
13
Q
What are the four types of noncovalent bonds?
A
Electrostatic Interactions
Van der Waals
Hydrophobic Interactions
Hydrogen bonding
14
Q
Describe the effects of hydrophobic interactions in an aqueous environment?
A
Water sequesters anions and cations
Nonpolar molecules are driven together
15
Q
Define hydrogen bond
A
Hydrogen atom shared by 2 atoms, usually an O or N
16
Q
Differentiate between hypertonic and hypotonic solutions
A
Hypertonic - water moves out of the cell and it shrinks
Hypotonic - water enters the cell and it swells and bursts
17
Q
What are the two basic equations needed to begin the derivation of the HH equation?
How do you derive the HH equation?
A
pH = -log[H+] Ka = [H+][A-] / [HA]
1) Ka= [H+][A-]/[HA]
2) Ka [HA]/[A-] = [H+]
3) -logKa -log[HA]/[A-] = -log[H+]
4) pH= pKA +p[A-]/[HA]
18
Q
What is the HH equation?
A
pH = pKa + log [A-]/[HA]
19
Q
Where is the buffering capacity of a system greatest?
A
pH = pKa, where [HA] = [A-]
20
Q
Where is the buffering capacity of a system mostly lost?
A
pH is one less or greater than pKa
21
Q
In a graph of OH- added by pH, where is pH=pKa?
A
Midpoint
22
Q
What does amphoteric mean?
A
Can act as either a base or an acid
23
Q
What are the three forms of an amino acid with a non-ionizable side group?
A
Both groups protonated → Zwitterionic form → Both deprotonated
24
Q
How many pKas will an amino acid have if it has an ionizable side group?
A
3
25
What is acidosis? Alkalosis?
Blood pH < 7.4 in acidosis, blood pH > 7.4 in alkalosis
26
What is the meaning of respiratory acid/alkalosis? Example?
Respiratory ___osis → a change in acid-base status induced by altered respiration
Hyperventilation lowers blood CO2 (alkalosis)
27
What is the meaning of metabolic acid/alkalosis? Example?
Metabolic ____osis → a change in acid-base status inducted by metabolic problems
Caused by excessive vomiting (alkalosis)
28
What is a domain of a protein?
Domains are two or more compact regions connected by a flexible segment
29
Define the secondary structure of a protein
Secondary structure is the spatial arrangement of amino acids that are close to one another
30
What amino acid forms disulfide bonds?
Cysteine, C
31
What is a protein cofactor?
Functional non-amino acid components
32
What are the two types of protein cofactors?
metals and coenzymes-small organic molecules
33
What are the two types of coenzymes?
Prosthetic Groups - Tightly bound coenzymes covalently bonded to the protein
Cosubstrates - Loosely bound coenzymes that dissociate and re-associate
34
What are the ten essential amino acids?
PVT TIM HALL
Phenylalanine, Valine, Threonine, Tryptophan, Isoleucine, Methionine, Histidine, Arginine, Leucine, Lysine
***Does not follow single letter symbols for the AAs
35
Define chirality
Chiral → the rotated molecule cannot be superimposed on mirror image
36
What is the chirality of the amino acids?
All except glycine are chiral, all are L, all except cysteine are S
37
What is the isoelectric point of an amino acid? How is it found?
The Ph where the net charge on AA is zero
| Average of the pKas when the AA is +/- 1
38
Which amino acids have aromatic rings?
Phenylalanine
Tryptophan
Tyrosine
39
Which amino acids have non-aromatic rings?
Proline
| Histidine
40
Which amino acids contain sulfur?
Cysteine
| Methionine
41
Which amino acids are negatively charged?
Aspartate
| Glutamate
42
Which amino acids are positively charged?
Lysine
Arginine
Histidine
43
What are peptides?
Small condensation products of amino acids
44
How are peptides named?
Naming begins on the N-terminus side and ends at the carboxyl-terminus
Using either the partial amino acid name with (yl) in between, the three letter code abbreviation with dashes in between, or the single letters with nothing in between
45
Describe the peptide bond. Is it flexible? Cis?
The bond between the carbonyl carbon of one amino acid and the nitrogen of the next has partial double-bond character, so it is rigid
Almost all peptide bonds occur in the trans configuration due to steric clashes from R groups
46
On a peptide, what are the phi angles?
Phi (𝝓) angles are between the 𝞪 carbon and the nitrogen of the amide
47
On a peptide, what are the psi angles?
Psi (𝜳) angles are between the 𝞪 carbon and the carbon of the carbonyl
48
What is a Ramachandran diagram for?
it shows favorable conformations of the phi and psi angles
49
What stabilizes an alpha helix? Where are the bonds?
Stabilized by H-bonds
CO of aa(n) bonds with NH of aa(n+4)
All amino acids except the ends are hydrogen bonded
50
How many amino acid residues are there for each turn of the alpha helix?
3.6 aa residues per turn of helix
51
Are alpha helices mostly left-handed, right-handed, or found equally in both? Why?
Right-handed due to steric clash
52
What are some amino acids that favor alpha helices?
Amino Acids → Ala, Glu, Leu
53
What stabilizes a beta sheet?
h-bonds
54
What differentiates the two forms of beta sheets?
Parallel → Each amino acid connects with two others
Antiparallel → Each amino acid connects with one other
55
Which amino acids favor beta sheets?
Amino Acids → Val, Ile, Thr
56
What bond promotes the formation of a reverse turn?
CO of aa(n) is H-bonded to the NH of aa(n+3)
57
What amino acid is commonly found in reverse turns and why?
Often include proline, especially in 𝜷 sheets due to low rotation around 𝞪 carbon
58
What is the structure of a coiled-coil protein?
2 right-handed 𝞪 helices wrap around each other to form a left-handed 𝞪-helical coiled coil
Central region of 300 aas with imperfect repeats of 7 aas
59
What bonds are involved in a coiled-coil?
Helixes are bonded by van der Waals, ionic, and disulfide cross-links
Number of disulfide cross links defines properties
60
How many residues are there per turn of a coiled-coil?
3.5 aa residues per turn
61
What amino acid is commonly found in coiled-coils and why?
Include leucine b/c hydrophobic
62
What is the composition of a collagen strand?
Regular aa sequence: every third is Gly, proline and hydroxyproline are also abundant
Gly is used due to small size, forms inside of superhelix
63
What is the structure of collagen?
3 strands combine to form a superhelical cable with 3 aas per turn
64
What stabilizes collagen?
Stabilized by steric repulsion of pyrrolidine rings and hydrogen bonds
65
What is the tertiary structure of a water-soluble protein?
Water-soluble proteins fold into compact structures with nonpolar cores
66
What is the tertiary structure of a trans-membrane protein?
Trans-membrane proteins have the opposite structures with polar centers
67
How do prion diseases spread?
a misfolded protein serves as a template for others
68
What are intrinsically unstructured proteins (IUPs)?
without a defined structure when not interacting with other proteins
69
What are metamorphic proteins?
exist in many conformations of about equal energy
70
What is amyloidosis?
improperly folded proteins result in protein aggregates (amyloid fibrils or plaques) due to the formation of large parallel 𝜷 sheets
71
What protein is involved in the following conditions: Alzheimer's, Parkinson's, and ALS?
Alzheimer's → Amyloid 𝜷 protein
Parkinson's → 𝞪-synuclein
ALS → superoxide dismutase enzyme (SOD)
72
What is specific activity?
ratio of enzyme activity to the amount of protein in the mixture
units per mg
73
What is yield?
(total) activity retained after each step
| %
74
What is purification level?
specific activity at that step over initial specific activity
75
What is the first step of the protein purification process?
Homogenate is centrifuged in multiple stages producing pellets of cellular materials and a supernatant consisting of the soluble portion of the cytoplasm
76
Describe gel-filtration or molecular exclusion chromatography
separates proteins on the basis of size using a glass column with porous beads, smaller beads enter beads so larger elute first
77
Describe ion-exchange chromatography
Separates proteins on the basis of charge using charged beads and solutions of salt or acid/base
Named based on charge of proteins eluted first
78
Describe affinity chromatography
Separates proteins on the basis of affinity for specific chemical groups
Wash with solution containing high levels of that chemical group
79
Describe high-performance liquid chromatography
A subtype of affinity chromatography with fine beads and high pressure
80
What is salting out?
Most proteins are less soluble at high salt concentrations, concentration depends on particular protein
81
What is dialysis?
Separate proteins from small molecules by semipermeable membranes with small pore sizes
Place a dialysis bag with a concentrated solution inside a buffer filled container. Smaller molecules and ions transverse the pores and emerge outside the bag
82
What is the general principle of gel electrophoresis?
Proteins migrate with a velocity that is proportional to the strength of the electric field and the charge on the protein and inversely proportional to the friction coefficient (mass, shape, medium)
83
Describe SDS-PAGE gels
Separate proteins by mass with the addition of SDS, which gives all proteins a high negative charge
Mobility is linearly proportional to the log of their mass
Polyacrylamide Gel is formed by combining acrylamide with the crosslinking agent MBAA
84
How can you visualize the results of your protein gel?
Coomassie blue or silver stains
| Radioactive labels
85
What is isoelectric focusing?
Gel with pH gradient, each protein moves until pH = pi
| Can be combined with SDS-PAGE
86
What factors influence sedimentation coefficient (s)?
Depends on the mass (of the particle), shape (of the particle), and density (both)
87
What is zonal/band/gradient centrifugation?
Used to separate proteins with different sedimentation coefficients
88
How does zonal/band/gradient centrifugation work?
Formation of sucrose gradient → addition of sample → swinging bucket rotor
89
Sanger's Method of sequencing polypeptides UPDATES
1-fluoro-2,4-dinitrobenzene binds to the N-terminus
Benzene with F, NO2, space, NO2
90
How do you determine amino acid composition (2-part)?
Heat at 110*C with 6 M HCl for 24 hours
Ion-exchange chromatography and ninhydrin indicator dye
91
Edmann's Method for N-terminus
Label N-terminus with phenyl isothiocyanate (PIT), treat with weak acidic conditions to remove the N-most acid
Benzene with N=C=S
92
Cleavage by cyanogen bromide (CNBr)
-COOH site of Met
93
Cleavage by trypsin
-COOH site of Lys, Arg
94
Cleavage by chymotrypsin
-COOH site of Phe, Tyr, Trp
95
Cleavage by pepsin
-NH site of Phe, Tyr, Trp
96
Removal of disulfide bonds for amino acid sequence determination
Reduce with thiols such as dithiothreitol (DTT) and beta-mercaptoethanol
97
Polyclonal antibodies
Polyclonal antibodies are derived from multiple antibody-producing cell populations
Heterogeneous mixtures of antibodies that are each specific for one epitope
98
Monoclonal antibodies
Monoclonal antibodies are derived from a single population, are identical, and respond to a single epitope
99
Antibody Production
Fuse normal antibody-producing cells with those with multiple myeloma
100
What does ELISA stand for?
Enzyme-linked immunosorbent assay
101
What is ELISA for?
Quantify the amount of protein present from enzyme-linked antibodies
102
Describe indirect ELISA
ANTIGEN-coated well → add ANTIBODY (binds to antigen)→ add enzyme-linked antibody (binds to first antibody) → add substrate (converted by enzyme into colored product that is proportional to the amount of SPECIFIC ANTIBODY)
test for HIV antibodies in blood
103
Describe sandwich ELISA
MONOCLONAL ANTIBODY-coated well → add ANTIGEN (binds to antibody)→ add enzyme-linked monoclonal antibody → add substrate (converted by enzyme into colored product that is proportional to the amount of ANTIGEN)
test for antigens in blood/urine, test for food allergens
104
Western Blotting
Proteins are separated by SDS, stained with an antibody, then with a second antibody connected to a chemiluminescent or fluorescent tag
105
Product and issue with mass spec
mass-to-charge ratio, conversion into gas phase
106
Compound that blocks the amino group of the incoming amino acid for peptide synthesis
t-Boc or tert-butyloxycarbonyl
107
Compound that facilitates peptide bond formation for peptide synthesis
DCC or dicyclohexylcarbodiimide
108
Product and issue with x-ray crystallography
3D structure, needs a crystalized molecule
109
Product and issue with NMR
3D structure, only some atoms show up
110
Product of cryo-electron microscopy
3D structure from many orientations
111
Condition with really stretchy skin
Ehlers-Danlos
