biochem: lec 2 pt.2

domain

• tert struc

• a globular cluster of 2* / supersecondary struc in peptide w/ >200 residues

subunit vs domain

Subunit = separate protein chain

Domain = region within a protein chain

what are the avg residues and diameter a domain has

• 40-200 residues

• 25 A

quat struc

• non-covalent connection btw 2 or more polypep chains

• ex: hemoglobin has 4 subunits → 4 poly pep chains

what’s the only covalent bond that can occur in quaternery struc?

disulfide bond (two cys)

what is the quat struc of transthyretin (homo-tetramer)

• 4 identical subunits

• 4 polypep

• 1 type of polypep

transthyretin tetramer & retinol binding protein

• protein-protein intxn

• TTR: tetramer, 4 polypep

• RBP: 1 polypep

• 1 TTR + 2 RBP -retinol

what do you get when you unravel TTR₄ + 2 RBP–retinol

2 types of poly pep chains

• 6 total poly pep

what’s the function of TTR binding RBP

• RBP transport retinol in blood,

  • too small and can get unfiltered by kidneys

• binding to larger TTR inc surface area & prevent from getting filtered

what are 2 advantages of quat struc

• can repair defects by replacing the flawed subunit

• provides structural base for activities (e.g allosteric/ cooperative binding)

fibrous protein

• long, elongated fibers/strands

• structural role over enzymatic role

• repeating 2* structures

  • ex: keratin, collagen

alpha-keratin

• strong, fibrous

• coiled coil 2* struc

• lots of Cys → gives sulfur smell when burns

  • present in hair, outer epidermis, horn, nails, feather

what is coiled coil

• the coiling of two alpha helices

• protofilament: pair of coiled coil (4 helices)

• filament: at least 4 protofibrils twisted

example of permanent hair wave (perm)

• hair before perm: S-S bind hold shape of keratin

• reducing agent (H): ammonium thioglycolate converts S-S —> SH + SH

  • makes hair flexible enough to change its shape

• oxidizing agent: hydrogen peroxide forms disulfide bonds

  • curling hair, new shape

collagen

• large amounts in vertebrate animals

• strong, insoluble fibers

• triple helix: 3 ind poly pep chains twist around e/o

• major component if connective tissues

  • bone, teeth, cartilage, tendon, and fibrous matrices of skin & blood vessels

structure of collagen

• side chains are covalently cross-linked via lysine/hydroxylysine residues

• cys is usually not present in collagen

collagen amino acid composition

• 30% gly

• 15-30% pro & 4-hydroxyprolyl (Hyp)

• others

converting proline to hydroxyproline

• prolyl hydroxylase (enzyme) and Vit C (coenzyme) adds OH group to C4

• tabilize collagen’s triple helix

scurvy

• collagen disease

• caused by absorbic acid (Vit C) deficency → poor collagen fibers

  • low activity of prolly hydroxylase

  • low of Hyp

  • weak collagen fibers

what does Hyp do but Pro can’t?

• Hbond

• the extra OH group form hbond —> provides stability to collagen’s triple helix

ways that enzymes bind to substrates

• geometric & electronic complementarity

• noncovalent forces (reversible binding)

• hydrophobic groups

• dashed lines (h-bond)

• “lock & key” and “induced fit”

protein are dynamic structures

• non-rigid, always move and change its shape —> important for their function

• binding can shift or change shape —> changes can occur in side chains, domains, or entire subunits.

what is the molecular weight of protein

• avg aa: 110 daltons

• avg protein: (# of aa) x 110

location of side chains depends on polarity

• nonpolar: inside, no contact w/ aqueous solvent

• charged polar: surface

• uncharged polar: can be out or inside

  • when buried inside → hbond w/ other groups

5 factors contribute to protein stability

• hydrophobic effect

• hbond

• ionic interactions

• disulfide bonds

• metal ions

what has the greatest influence on protein stability

• hydrophobic effect

• due to increase in entropy water moc

• major determinant of native protein struc

what are the central features of protein struc

• hbond

• minor contributions to protein stab

what happens to hbond when protein unfolds

• small diffrence in h-bond free energies

• unfolded forms h-bond w. water moc

ionic interaction in protein stab

• occurs btw oppo charged side chains

• ion pair/ salt bridge

  • formation makes side chains more organized —> loss in entropy

• minor contribution

  • ionic attraction is fav (stability) → loss of freedom is unfav (reduce stability) → overall effect is small

disulfide bonds in protein stab

• form btw proteins outside of the cell

• rare inside cell bc cytoplasm is a reducing env

metal ions in pro stab

• help stabilize small protein structural motifs

• ex: zinc finger

zinc finger

• 25-60 residues around one/two Zn 2+ ions

• aa side chains Cys & His hold zinc ions in place

• function: bind DNA, RNA, proteins

how does zinc binding contribute to stability?

small region of protein may not maintain its proper shape w/o zinc ion -> binding helps stablizing motif and perform its function

5 ways protein can denature

• heat

• acid/base

• detergents

• reducing agents

• chaotropic agents

heat in den

• proteins can unfold/ melt above 100 C

acid/base

• acidic: protonate & impact hbond/disulfide bridge

detergents

ex: sodium dodecyl sulfate (SDS)

• disrupts noncovalent intxn (hydrophobic, ionic)

• unfolds and give protein neg charge

reducing agents

• dithiothretiol, b-mercaptoethanol

• break disulfide bonds

chaotropic agents

• guanidinium ion, urea

• inc solubility of np stuff in water

• disrupt h-bond & hb intxn

Polyphenol oxidase (PPO) in fruits

• enzyme that causes browsing when cutting fruit

how to decrease browning in fruits

• inactivate PPO

  • heat (cooking): deactivate enzyme

  • lower pH on surface: lemon juice

  • lower O2 level: enzyme has no oxidizing power

    • put fruit in water/ vacuum

what happens when dopamine get oxidized

dopamine —> dopamine - o -quinone

• dopamine loses 2 e- & 2H+ rel

renaturing of protein

• urea denature protein & mercatoethanol cleaves disulfide bonds

  • 4 disulfide bonds —> 8 SH grps

• removal of reagents & O2 present allow protein to reature & reform

molecular chaperones

• enzymes

• help proteins fold at early stage

how do molecular chaperones help in protein folding

• bind to unfolded & partially folded poly pep chains

  • help fold to native struc

  • prevent exposed hp regions of unfolded proteins from aggregating

heat shock protein (Hsp)

• important chaperones

• help proteins maintain / regain stcuture

• at high temp: Hsp production inc. help by come around & hold protein (hug)

  • V shape that wraps around protein

Hsp 90 and cancer

• cancer cells can use Hsp90 to keep proteins supproing tumor grow and surviva

• blocking Hsp90 can break down these protein → harder for cancer cell to grow

• cancer drug target protein