biochem unit 1 post quiz 1

pka of amino group

10-12

pka of COO- group

2-3

for all amino acids besides proline and glycine, the alpha carbon is bonded to:

acidic carboxyl group

a basic amino group

hydrogen

unique side chain

chirality

cannot be superimposed on mirror image

all amino acids besides glycine are chiral

how do you determine L and D forms

COO- to top, R group to bottom, where is amino?

amino acids in natural proteins are in what configuration

L

where do D amino acids occur

small peptides in peptidoglycan of bacterial cell walls

peptide antibiotics

neurotransmitters (D-glutamate)

platypus venom

all three aromatic amino acids absorb UV light at

270-280 nm

side chains in what class of amino acids can form H bonds

polar, uncharged

where do cysteines form disulfide bridges with each other

extracellular environment to stabilize individual proteins and protein complexes

cytosol is what kind of environment

reducing

pka of lysine

10.53

basic, positively charged

pka of arginine

12.48

basic, positively charged

pka of histidine

6.00

basic, positively charged

pka of aspartate

3.65

negatively charged, acidic

can bind Mg, Ca, Mn, Zn, ionic interactions

pka of glutamate

4.25

negatively charged, acidic

can bind Mg, Ca, Mn, Zn, ionic interactions

what form of amino acids, carbs, and nucleic acids do living organisms use

L amino acids

D carbs

D nucleic acids

ionization of amino acids

-amino acids are weak polyprotic acids, each amino acid has at least two titratable groups

-at acidic pH, carboxyl group is protonated and the amino acid is in the cationic form

-at neutral pH, carboxyl group is deprotonated but the amino group is protonated. the net charge is zero, zwitterion

-at alkaline pH, the amino acid group is neutral -NH2 and the amino acid is in the anionic form

diprotic amino acid

pk1- carboxyl group

pk2- amino group

isoelectric point of amino acid

pH where there is an equal amount of positive and negative charges

isoelectric point for glycine

5.97

In his, the imidazole side chain also contributes a

titratable group

therefore is triprotic

if pH is greater than pka

deprotonated

if pH is less than pka

protonated

what is the only AA that can be an effective buffer at physiological pH

histidine

its R group has a pka of 6

no other AA side chain has a pka near neutral pH

peptide bonds

peptides form covalent bonds between the alpha-COOH and alpha-NH2 groups of two amino acids, resulting in loss of water molecule

O=C-N-H

peptide definition

two or more amino acids joined covalently by a peptide bond

polypeptide definition

many AA joined together by peptide bonds (MW less than 10,000) (~15-50 AA)

protein definition

macromolecule with one or more polypeptide chains (50+ AA)

numbering and naming starts from which terminus

N

other relevant small peptides

oxytocin (9 AA)- stimulates uterine contractions

Bradykinin (9 AA)- causes tissue inflammation

Polypeptide examples

insulin- pancreatic hormone, needed for sugar metabolism, 2 polypeptide chains (30 AA and 21 AA)- not a dipeptide

Glucagon- pancreatic hormone, opposes insulin (29 AA)

average sized protein

hemoglobin with 574 AA and MW of 64,500

protein sequence logos

graphical representations of patterns within a multiple sequence alignment

provide a richer and more precise description of sequence similarity than consensus sequences and can rapidly reveal significant features of the alignment otherwise difficult to perceive

height of stack indicates sequence conservation at that position, measured in bits

height of symbols within stack reflect relative frequency of corresponding amino or nucleic acid at that position

native fold

specific three-dimensional conformation that enables a protein to fulfill a specific biological function

protein conformation is stabilized by

disulfide bonds

weak, non covalent interactions

hydrophobic effect relating to protein folding

release of water molecules from the structured solvation layer around the molecule as protein folds increases the net entropy

correctly position hydrophobic side chains depending on environment

hydrogen bonds relating to protein folding

interaction of N-H and C=O of the peptide bond leads to local regular structures such as alpha helices and beta sheets

side chain side chain interactions

van der waals relating to protein folding

weak attraction between all atoms contributes significantly to the stability in the interior of the protein

electrostatic interactions/ionic bonds relating to protein folding

stronger interactions between permanently charged groups

salt bridges especially buried in the hydrophobic environment strongly stabilizes protein

primary structure

amino acid sequence, includes all covalent bonds between amino acids and locations of disulfide bonds

spatial arrangement is unspecified (high degree of flexibility and rotation about bonds, except peptide)

clearly distinguishes one protein from another

amino acid sequence and protein function are closely linked, but difficult to tell function at this level

sickle cell mutation

single AA substitution

abnormal hemoglobins crystallize, deforming red blood cells and leading to clogs in tiny blood vessels

paralogs

same species

orthologs

different species

secondary structure

regular, recurring arrangements in space of adjacent amino acid residues in a polypeptide chain

most prominent secondary structures are alpha helices (coils) and beta sheets (folds)

irregular arrangement of the polypeptide chain is called a random coil

any one protein can be all one type of secondary structure, or a mixture

peptide bond rigidity

rigid and planar constraining the protein to certain allowed conformations

no rotation around peptide bonds BUT

rotation around C alpha bonds is permitted

phi bond

Calpha-amide nitrogen

psi

Calpha to carbonyl carbon

Ramachandran plot

shows common secondary structural elements and the acceptable range of rotation

the alpha helix

-helical backbone is held together by hydrogen bonds between the backbone amide of an “n” and carbonyl group of the n+4 amino acid

-right handed helix with 3.6 residues (5.6 A*) per turn

-hydrogen bonds are aligned roughly parallel with the helical axis

-side chains point out and are roughly perpendicular with the helical axis, H bonds are mostly parallel with helical axis

-residues 1 and 8 align on top of each other, heptad repeat

-the inner diameter of the helix (no side chains) is about 4-5 A*

-the outer diameter of the helix (with side chains) is 10-12 A*, fits well into the major groove of dsDNA

-some amphipathic alpha helices can form coiled coil dimers

in an alpha helix, where are hydrophobic AA found?

in A and D positions

primary sequence affects helix stability

small hydrophobic residues such as Ala and Leu are strong helix formers

pro acts as a helix breaker because the rotation around the N-alphaC bond is impossible

gly acts as a helix breaker because the tiny R group supports other conformations, too flexible

attractive or repulsive interactions between side chains 3-4 AA apart will affect formation

residues near N and C ends don’t necessarily H bond like the rest, but they can stabilize the helix (helix capping)

beta sheets

sheet-like arrangement of the backbone is held together by hydrogen bonds between the backbone amides and carbonyl groups in different strands

side chains protrude from the sheet alternating in up and down direction

which AA will or will not be found in beta sheets

found: large, aromatic (Tyr, Phe, Trp) branched (Leu, Val, Ile)

not found: Gly, Pro

parallel beta sheets

in parallel beta sheets the H bonded strands run in the same direction

results in bent, weaker H bonds

individual strands can be close, or distant, in the primary structure

antiparallel beta sheets

H bonded strands run in opposite directions

resulting in stronger, linear H bonds

individual strands can be close, or distant, in the primary structure

ribbon diagrams of secondary structure

most proteins are globular in shape as a result of frequent turns or loops in the polypeptide chain that connects beta strands and alpha helices

strands, and sometimes helices, will have arrows to indicate N and C termini of the secondary structural element

beta turns help organize secondary structures

beta turns occur frequently whenever strands in beta sheets change direction

the 180 degree turn is accomplished over four amino acids

the turn is stabilized by a hydrogen bond from a carbonyl oxygen to amide proton three residues down the sequence.

common AA in beta turns

proline in position 2 or glycine in position 3

proline in cis conformation forms a tight turn, most peptide bonds are a trans conformation

tertiary structure

overall 3D arrangement of all atoms in a protein

includes long range contacts between AAs in a single polypeptide chain

stabilized by numerous weak interactions between AA side chains (largely hydrophobic and polar interactions, can be stabilized by disulfide bonds, side chain with backbone interactions are also possible)

major classes of tertiary structure

fibrous, globular, membrane, intrinsically disordered

fibrous proteins

often structural rather than dynamic and are water insoluble

typically contain high proportions of alpha helices (keratin) or beta pleated sheets (silk fibroin)

underlying structure is relatively simplistic

high proportion of hydrophobic AA

extensive supramolecular complexes

secondary structures and properties of fibrous proteins- alpha helix

tough, insoluble protective structures of varying hardness and flexibility

e.g. alpha keratin of hair, feathers, and nails

secondary structures and properties of fibrous proteins- beta conformation

soft, flexible filaments

e.g. silk fibroin

secondary structures and properties of fibrous proteins- collagen triple helix

high tensile strength, without stretch

e.g. collagen of tendons, bone matrix

alpha keratins

typical right handed alpha helices

super twisting helices, in a left handed fashion, wrap around each other to make a very tight coiled coil

cross linked by disulfide bonds

real life example using alpha keratins

permanent waving of hair

reduce disulfide bonds

moist heat breaks H bonds and causes uncoiling of alpha helix

remove reducing agent, add oxidizing agent, new S-S bonds

collagen

provides strength and structure

connective tissue (tendons, cartilage, organic matrix of bone, cornea)

each protein folds into left handed helix (alpha chains, NOT alpha helix)

coiled coil of three separate alpha chains supertwisted around each other in a right handed manner to provide strength (like a rope)

covalent cross links

globular proteins

enzymes, transport proteins, motor proteins, regulatory proteins, immunoglobins, etc

tight spherical structure, hydrophobic to middle

alpha helices, beta sheets, beta turns, etc. can all be found

arrangement of different secondary structural elements

• compact conformation

• folding provides structural diversity

general rules

• bury nonpolar AA R groups

• distant segments may come together, but not the norm

• optimize number of weak interactions

  • 
    

globular proteins: motifs

specific arrangement of several secondary structure elements

• all alpha helix

• all beta sheet

• combination

sometimes considered supersecondary structures

motifs can be found as reoccurring structures in numerous proteins

proteins are made of different motifs folded together

weird globular protein motifs

not secondary structures in themselves

twisted beta sheet

alpha alpha corner

beta alpha beta loop

beta barrel

globular proteins: domains

can have multiple domains on one protein, can have motifs in domains, domains by themselves are stable

-a domain is a relatively stable, independently folded region within the tertiary structure of a globular protein

-each domain may encompass one or more motifs

-domains having more than 30% of their AA seq in common normally adapt same folding pattern

-a single protein can have several domains with each domain performing a different function (enzyme, docking, regulatory, etc)

quaternary structure

results from the association of two or more polypeptide subunits into a larger functional cluster

side chain-side chain and side chain-polypeptide backbone interactions drive these associations. can have stabilizing H bonds

collagen is a fibrous protein of three polypeptides that are supercoiled like a rope

hemoglobin is a globular protein with two copies of two kinds of polypeptides

what drives quaternary association?

stability: reductio of surface to volume ratio

genetic economy and efficiency

bringing catalytic sites together

cooperativity

• biological function may be regulated by complex interactions of multiple subunits

how can proteins be denatured

strong acid or base, organic solvents (more nonpolar than water), detergents, reducing agents, salt concentration, heavy metal ions, temperature, mechanical stress

denatured protein (inactive form)

lost enough structure, can no longer function. does not have to go all the way back to primary

denaturation does not disrupt primary structure

Levinthal’s paradox

it is mathematically impossible for protein folding to occur by randomly trying every conformation until the lowest energy one is found.

search for the minimum is not random because the direction toward the native structure is thermodynamically most favorable

nonpolar inside, polar outside

assistance to ensure proteins fold properly

small polypeptides (<100 AA) often form with no intermediates

larger polypeptides often require several intermediates (molten globules)

chaperones

ribosome brings partially folded or misfolded protein to GroEL barrel, ATP and GroES cap join, ADP + P, GroES cap, and properly folded protein emerge

how do chaperones assist protein folding

prevent inappropriate protein-protein interactions

provide a more isolated environment

help folding occur rapidly and precisely

two major classes: Hsp70s Hsp60s (chaperonins)

PDI- protein disulfide isomerase

reduces disulfide bonds

PPI- peptide protyl cis-trans isomerase

moves from cis to trans if needed

intrinsically disordered proteins

composed of AA whose higher concentration forces less-defined structure: Lys, Arg, Glu, Pro (all charged, no hydrophobic effect)

NPC is one example

overview of globular protein functions

storage of ions and molecules- myoglobin, ferritin

transport of ions and molecules- hemoglobin, serotonin transporter

defense against pathogens- antibodies, cytokines

muscle contraction- actin, myosin

biological catalysis- chymotrypsin, lysozyme

protein function: binding to ligands

function relies on interactions with other molecules

binding of molecules to proteins is reversible

ligand: any molecule that a protein can bind to in a reversible manner; often a small molecule

binding site: site on protein where ligand binds, specific

ligand binds via same noncovalent forces that dictate protein structure- allows interactions to be transient

reversible binding of ligands is essential

specificity of ligands and binding sites, high specificity, only certain ligands bind

ligand binding is often coupled to conformational changes, sometimes quite dramatic (induced fit)

in multi subunit proteins, conformational changes in one subunit can affect the others (cooperativity)

interactions can be regulated

high specificity can be explained by the complementarity of binding site and ligand

size, shape, charge, hydrophobicity

lock and key model- rigid interaction

induced fit- both protein and ligand can change conformations upon binding, makes binding site more complementary to ligand

myoglobin

compact globular protein composed of a single polypeptide chain 153 AA in length

carries and stores oxygen for muscles

contains a heme prosthetic group- a porphyrin ring complexed with iron ion

oxygen binds to Mb via the heme as amino acids alone cannot bind O2

porphyrin ring- heme

6 coordination sites: 1-4 by Ns, 5=AA, 6=O2

hemes are also used in ETC