Topic 4 Protein structure and function

Proteins

main building blocks of the cell

Protein functions

enzymes, structural proteins, transport, motor proteins, storage (of left over AA), signalling, receptors, transcription regulators

Polypeptide backbone

repeating, core atoms (N-C-C)

N-terminus

end that carries an amino group

C-terminus

end that carries a carboxy group

Side-chain (r-group)

gives amino acid its identity and unique properties (charege? polar/nonpolar, chemically reactive?)

polypeptide chains

flexible rotaion around singl bond, shape constrained by weak interactions (backbone/r groups), polar amino acid around outside, nonpolar side chains forced together

Hydrophobic force

nonpolar side chain forced together towards inside of folded protein

conformation

shape of polypeptide, final folded structure, energetically favourable

denaturation

protein loses it conformation, caused by disruption of non covalent bonds, cause by heat or pH change

renaturation

spontaneous of refolding of protein when proper condition provided

chaperone proteins

Assist polypeptide to fold into most energetically favourble conformation

isolation chamber

prevent polypeptide from aggrgating with other polypeptides, type of charperone protein

Protein length and shapes

can be 30-10000 AA, averages 50-2000 AA, filament/sheet/sphere/rings

types of protein models

backbone, ribbon, wire, space filling

backbone model

show shape, doesn’t show sidechain (only backbone)

ribbon model

shows folding patterns/secondary structure (alpha/beta)

wire model

show back bone and side chain

space filling model

most accurate, space that AAs take up

primary structure

Amino acid sequence

secondary structure

alpha helices, beta sheets

tertiary structure

full 3D confromation, some proteins stop here

quaternary structure

multiple polypeptides interacting to form protein

alpha helix

secondary structure, reight-handed helix (complete turn every 3.6AA), H bond between every 4th AA, abundant in embedded cell membranes, many similar subunit in repeated relationship

Coiled-coil

2-3 alpha helices wrapped aroun eachother, very stable, elongated proteins, keratin and myosin

Beta-sheets

secondary structure, rigid structure at core, H bond between neighbouring segments, parallel/antiparallel

parallel Beta-sheets

run in the same direction

antiparallel beta sheets

run in opposite direction

Amyloid structure

Beta-sheets stack together with interdigiated side chain, storage of peptides or protein hormones, misfolded protein form damaging amyloid

Prions diseases

Scapie (sheep), Bovine spongiform (pig), encephalopathy (mad cow), Creutzfeldt-Jakob’s (human), chronic wasting (deer)

prions

infectious(move other cells/inbetween species) misfolded proteins (cause other to misfold) which contain amylod structures

amyloid that damage cells

Alzhimer, PArkinson, Huntington

Protein domain

segment of polypeptide that can fold independently into compact, stable structure

unstructure sequences

short polypeptide chains, links domains together

Protein families

group of proteins that closely resemble eachother, each has own distinct enzymatic function

binding sites

regions on protein’s surface that interacts (noncovalently) with another moelcule

Subunit

each polypeptide chain, could have multiple domain

Dimer

2 identical polypeptide chains bound together

tetramer

4 subunits bound together

Globus proteins

chains of identical proteins (often helix), cage-like spherical shells, mixtures of various proteins and RNA/DNA

Fibrous proteins

alpha keratin, intermediate filaments, extracellular matrix

identical proteins (often shape of helix)

actin filaments, microtubules

Cage like spherical shell

capsids (protein coat of virus)

mixture of various proteins and RNA/DNA

ribosomes, viruses

alpha-keratin

fibrous protein, dimer of 2 identical subunit, colied coil, extremely stable (long live), hairs/horns/nails

intermediate filaments

fibrous protein, rope-like, gives cell mechanical strength

Collagen

fibrous protein, extracellualr matrix, 3 peptide in hexli, glycin at every 3rd position at core, collagen fibrils (overlapping array)

Elastin

fibrous protein, extracellualr matrix, loose unstructured covalently linked elastic meshwork, enables to stretch without tearing (skin/arteries/lung)

stable covalent cross linkages

tie amino aicd in same chain or larger complexes, Disulfide bridges, only proteins being excreted

Disulfide bridges

link -SH groups from cystein side chains, don’t form in cytosol

protein binding

proteins bind other molecules, tight and long lived or weak and short lived

examples of protein binding

antibodies to virus/bacteria/WBC, enzymes to substrates, actin bind eachother (filament)

ligand

substance bound by protein, weak bonds between

binding site

region of protein ligand associates, cavity on protein surface, regulates protein activity, can use to attach to location in cell

substrate

ligand that binds enzyme

active site

binding site in enzyme

transition state

conformation of enzyme-substrate complex, lowers activation energy

metabolic pathway

network of enzymatic reactions, product of one is reactant of next

cofactors

small inorganic molecules that aid enzymes

coenzymes

small organic molecule that aid enzymes, from vitamins in diet

examples of cofactor

iron in heme groups, zinc in carboxypeptidase

examples of coenzymes

biotin transfer carboxyl groups, retinal in rhodopsin (absorb light)

Control of protein

gene expression, rate of protein degradation

how protein activity controlled

confine sub-celluar compartments, adjust activity using regulartory sites

regulatory sites

sites where molecules bind, alter rate at which enzymes functions, usually cause conformational change

Feedback inhibition

enzyme early in pathway is affected by molecule produced later

negative regulation

later product prevents earlier enzyme from acting

positive regulation

product in one branch stimulates enzyme in another

allosteric proteins

2 different conformations(active/inactive), spontaneuosly switch between 2 until ligand stabilizes in correct conformation

phosphorylation

attach phosphate to amino acid (serine) side change, negative chargecauses conformational change, removal returns to original

Protein kinase

transfer phosphate from ATP to ser -OH

protein phosphatase

removes phosphate group

Protein modifications

phosphorylation to ser, acetyl to lysine (histones), Fatty acids to cysteine (membrane proteins), Ubiquitin (degradation)

GTP binding proteins

active conformation when GTP bound, hydrolyzation (to GDP) causes inactive conformation, reactivation stimulated by cell signals

Motor proteins

generates forces responsible for muscle contractions and celluar movement, unidirectional conformation changes

unidirectional conformational change

one step is irreversible, ATP is hydrolysis (release free energy)

protein machines

highly coordinated linked set of proteins, hydrolysis of ATP/GTP drives ordered series of conformational change, successive reactions in series

Scaffold proteins

large molcules, contian binding sites recognized by many proteins, enhance rate of cell process by confining to one area, rigid/structure or elastic/unstructured, could be RNA

Biomolecular condensates

collection of proteins and RNA, held by continuously shifting weak interactions, fluid membraneless subcompartment that perform function, contains scaffold, non-covalently bonded, separate from surrounding

clients

molecules become concentrated on scaffold

cell homogenate or extract

content released from broken cell

Fractionation

separates class of molecules, by centrifugation

centrifugation

used for fractionation, separate chunks and unbroken cells

ultra centrifugation 

used for fractionation, separates membrane

chromatography

separates individul components into fractions, based on properties of protein

affinity chromatography

separates polypeptides, based ability to bind 

electrophoresis

polypeptides migrate through gel, different speeds depending size and net charge

Mass spectroscopy

determines mass of peptide fragment, identification from database, apply known genetic code

mass spectroscopy mode of action

blast peptide with laser, electrically charged(all group ionized) and gaseous, detector relates time to reach to mass and charge 

X-ray crystallography

take foled protein, fix in matrix to crystalize, x-ray bounce off based electeron density, use detector to read, use to give detail picture/data on shape

Nuclear magnetic resonance (NMR) spectroscopy

use magnets to pull/push functional groups, protein can’t be to big, use to give detail picture/data on shape

Cryo-electron microscopy (cryoEM)

transmission electron microscopy, take 2D image every side and compile, fix folded protein in ethanol ice, use to give detail picture/data on shape

why knowing protein structure important

human health - design new drugs to alter metabolic pathway/ stop infections, basic understanding how cell operates

sequence patterns

same domain over and over predict plays same role in all proteins bcprotein famillies

AI in protein structure discovery

predict structure high accuracy, accelerates pace of biological research

Genetic engineering functions

mass-produce therapeutic drugs, New proteins and enzymes (modified known) that perfrom unusual task, targetted drug delivery