ALL THE THINGS Flashcards
What is the strongest type of molecular bond?
Covalent
What are the other electrostatic bonds seen when analyzing proteins?
Ionic bonds, hydrogen bonds, van der Waals
What other interactions are commonly seen when looking at proteins?
hydrophobic interactions
Noncovalent interactions between specific amino acids determine what?
the folded shape of the protein
How do nonpolar groups hide from water? What does this due?
cluster together; decreases the disruption of H-bonds with water solvent; more energetically stable
What does a hydropathy plot measure?
hydrophobicity in a protein
What determines the chemical properties of the amino acid?
the nature of the R group
A
Alanine, Ala
R
Arginine, Arg
N
Asparagine, Asn
D
Aspartic Acid, Asp
C
Cysteine, Cys
Q
Glutamine, Gln
E
Glutamic Acid, Glu
G
Glycine, Gly
H
Histidine, His
I
Isoleucine, Ile
L
Leucine, Leu
K
Lysine, Lys
M
Methionine, Met
F
Phenylalanine, Phe
S
Serine, Ser
T
Threonine, Thr
W
Tryptophan, Trp
Y
Tyrosine, Tyr
V
Valine, Val
P
Proline, Pro
What are the essential amino acids that are nutrional requirements?
R, H, I, L, T, K, M, F, W, V
Which amino acids have nonpolar, aliphatic R groups?
G, A, P, V, L, I, M, C
C behaves like it is hydrophobic despite having a reactive R-SH group
What is special about Glycine?
it’s small R-H group confers flexibility in proteins
What does Proline’s unusual ring structure cause in protein structure?
Kinks
Which amino acids have aromatic R-groups?
F, Y, W
Which of the aromatic R group amino acids is not hydrophobic?
Y
Which of the aromatic amino acids absorbs the highest wavelength of UV light?
W with 280nm; F only at 260 nm; nucleic acids at 260 nm
At what absorbance level can proteins be quantified using UV light?
280 nm
Which amino acids have a polar, uncharged R group?
S, T, C, N, Q
Which amino acids have positively charged (basic) R groups?
K, R, H
Which amino acid have negatively charged (acidic) R groups?
D, E
By what reaction are peptide bonds formed and is it a spontaneous reaction?
condensation (dehydration) and NO!
The peptide bonds is ___ , ____ , ____ . There is no rotation at _____ _____ .
rigid, planar, has dipoles.
Peptide Bonds
What is the primary structure of an Amino Acid?
the linear sequence of amino acids in the polypeptide chain
In secondary protein structure, local regions of amino acids are folded into a limited set of what distinct structures?
alpha helix, beta sheet, and beta turn
What does the secondary structure generally involve in terms of bonds?
hydrogen bonds between carbonyl and amide groupsin the peptide bonds
Are R groups involved in secondary structure?
NO
What do alpha helices look like and how many amino acids per turn?
rigid, right handed helix formin a rod-like structure with 3.6 amino acids per turn
How many residues apart are the hydrogen bonds for alpha helices?
4
What if proline is present in an alpha helix?
proline is a helix breaker; it destablizes the helix;
also broken by bulky or charge R groups
Why is G not present in alpha helices?
confers too much flexibility to the helix
How are peptide chains arranged to form beta pleated sheets?
parallel or antiparallel
Where are hydrogen bonds formed in beta sheets?
between adjacent peptide chains
How are R groups positioned in beat pleated sheets?
alternate above and below the plane
What angle and how many amino acids are involved in Beta/Reverse turns?
180 degree turn of the peptide chain. 4 amino acids usually P and G
What is the tertiary structure of proteins?
overall 3D arrangement of the entire peptide chain
spatial organization of structures
What does the tertiary structure determine?
function of the protein
alterations due to mutations can abolish or alter functions
What is the quaternary structure of proteins?
spatial arrangement of polypeptide chains in a multi chain protein complex and the nature of their contacts
What does the quaternary structure determine?
function of the protein complex
How are tertiary and quarternary structures stabilized?
noncovaelent interations between the peptide bond and R-groups. also done by disulfide bonds
What are protein domains?
stable arrangements of several elements of secondary and tertiary structure
What do separate domains confer?
separate functions
Small proteins/large proteins have how many domains and functions?
Small: one domain, one or few functions
Large: multiple domains, multiple functions
How do domains act as evolutionary units?
copying and shuffling of domains led to formation of many protein families
What are 3D folds of protein structures associated with?
specific activity; e.g. ATP binding
What is used to analyze amino acid sequence alignment?
BLAST algorithm
What can the BLAST algorithm be used to identify?
homologous proteins from a sequence database
What are Orthologs?
genes in different species, evovled from a common ancestor gene; same function in the different species
What are Paralogs?
imperfect copies of genes within species (members of gene families); generally different but likely related functions
What is Column Chromatography?
separation of a crude protein sample in a buffer (mobile phase) through a stationary porous matrix
stationary phase properties and it’s interaction with the proteins form the basis of the separation
How are proteins separated in Column Chromatography?
separates based on some property of the protein
How are proteins purified in Column Chromatography?
sequential column chromatographies with different stationary phases
What is another name for Size Exclusion Chromatography?
Gel Filtration Chromatography
On what basis are proteins separated in Size Exclusion Chromatography?
separation on the basis of size
What is the solid phase size exclusion chromatography?
a porous bead
Do small or large molecules move fast or slow in Size Exclusion Chromatography?
Small molecules enter the beads and migrate slowly
Large molecules less likely to enter the beads, migrate more quickly
Which proteins are the first to elute in Size Exclusion Chromatography?
largest proteins
How do proteins separate in Ion Exchange Chromatography?
on the basis of charge
Which proteins migrate slowly in ion exchange chromatography?
proteins with opposite charge bind the resin and migrate slowly, unlike proteins that have the same charge as the resin
How do proteins separate in affinity chromatography?
on the basis of binding properties
how efficiently they bind a given ligand
Which proteins are retained in affinity chromatography?
only proteins that bind the ligand are retained; can elute because of competition with free ligand
What is Gel Electrophoresis?
separation of molecules in an electric field through a porous gel
What 3 things affect the rate of migration of molecules in Gel Electrophoresis?
size, conformation, and charge
How are molecules separated in SDS-PAGE?
on the basis of size
requires proteins to be unfolded without disulfide bonds
Which proteins migrate more quickly in SDS-PAGE?
smaller proteins migrate more quickly
opposite to size exclusion chromatography
What can be estimated from SDS-PAGE?
estimate the molecular weight of a protein
What is the isoelectric point of a protein?
pH at which there is no net charge
How are proteins separated in Isoelectric Focusing?
on the basis of their isoelectric points
How is the pH gradient established in isoelectric focusing?
by using Ampholytes in a gel
How long do proteins migrate in isoelectric focusing?
until they reach their isoelectric points
What technique is the best for separating complex protein mixtures?
2D Gel Electrophoresis
How are proteins separated in 2D gel electrophoresis?
separated through a gel in 2 perpendicular directions using a different basis of separation in each
What are the two binding constants?
Koff = dissociation rate
Kon = association rate
What is the equation for dissociation rate?
dissociation rate = koff [AB]
What is the equation for association rate?
association rate = kon [A] [B]
How do you solve for the equilibrium constant?
[AB]/[A][B} = kon/koff = K = equilibrium constant
What is the equilibrium constant (K) usually called and what is it’s inverse?
Ka (M^-1) and the inverse is Kd (M)
The stronger the binding between ligand and receptor —
the higher the association constant (Ka)
the lower the dissociation constant (Kd)
