Chapter 6: protein structure pt. 2 and 3

Question 1

Protein can be denatured by…

Answer 1

1. heating
2. Chaotropic agents

Question 2

proteasome

Answer 2

proteolyzed or refolds misfolded proteins

*only some proteins can be refolded or renatured

Question 3

chaotropic agents

Answer 3

disrupt interactions that stabilizes tertiary structure

Question 4

Christian Anfinsen

Answer 4

1. Used the correct folded state of RNase A
2. denatured RNase A by using urea and beta-mercaptoethanol (reduced disulfide bonds)
3. dialyzed out urea and most beta-mercaptoethanol to catalyze formation of correct disulfide bonds
4. the RNase regained activity

-showed that primary structures determines tertiary structure
*-delta G=when its in its native state

Question 5

Energetics of protein folding

Answer 5

the net delta G is negative=slightly favored

-delta G fold = -0.4 KJ/mol AA
-100 AA = -40 KJ/mol
*net energy of stabilization is small

Question 6

Levinthal’s paradox

Answer 6

-proteins DONT sample all possible folding conformations
-fold by specific pathway

*most proteins fold in less than 10 secs

Question 7

model of protein folding

Answer 7

1. the polypeptide collapses in upon itself due to the hydrophobic effect
2. secondary structure forms
   *molten globule state: lot of secondary structure and little tertiary structure
3. tertiary structure begins to form
4. then final native state forms

*takes less than 10 secs

Question 8

Folding Funnel

Answer 8

energy landscape guides protein towards native structure

Question 9

Folding in Vitro

Answer 9

-easy to fold
-folding by dilution in buffer

Question 10

Folding in vivo

Answer 10

-difficult to fold
-the cellular environment is crowded so there is a high potential for misfolding and precipitation (aggregation)

Question 11

Chaperons

Answer 11

-proteins that help other proteins fold in vivo
-also called Heat shock proteins (HSPs) bc they were discovered during HS

*most proteins can fold without chaperons

Question 12

DnaJ and DnaK

Answer 12

-prevents precipitation/aggregation
-assists folding
-coats the unfolded protein and preventing denaturing/precipitation

Question 13

GroEL and GroES: info

Answer 13

-also called Hsp60
-large chaperones called chaperonins
-actively fold proteins in prokaryotes, cytosol of eukaryotes, and mitochondria

-GroES: cap that seals unfolded protein from the outside
-GroEL: barrel

Question 14

GroEL and GroES: mechanism

Answer 14

1. protein in the barrel
2. cap goes from one side to the other side of barrel?
3. then it caps the end of the barrel that the protein went in
4. completely folded protein comes out of the barrel

*requires energy from ATP hydrolysis

Question 15

Protein disulfide isomerase

Answer 15

facilitates formation of correct disulfide bridges

Question 16

peptidyl proline isomerase

Answer 16

catalyses cis-trans isomerisation of peptide bonds involving proline

Question 17

How do protein folding diseases occur

Answer 17

-usually misfolded proteins get refolded immediately by a proteasome but sometimes they can accumulate as precipitates which can result in severe, chronic degenerative diseases that affect the brain and CNS

-Disease often occurs when protein with alpha-helix conformation misfolds into a precipitating beta-sheet conformation
*form a common cross-beta helical core filament structure

-leads to aggregation into amyloid fibrils masses which can form amyloid deposits (amyloid plaque) in brain
*can cause neuronal apoptosis

Question 18

Precipitated proteins are associated with what diseases?

Answer 18

1.Prions related illnesses
- like Transmissible Spongiform
Encephalopathies (TSE’s)
-Bovine Spongiform
Encephalopathy (BSE)/ mad cow’s
disease
- Creutzfeldt-Jakob disease (CJD,
human BSE)
2. Amyloid related illnesses
- Alzheimer’s,
-Parkinson’s
-ALS

Question 19

What is misfolding caused by

Answer 19

-genetics: through mutations
-transmission: Mad cow disease

Question 20

protein folding mechanism

Answer 20

1. native monomer
2. misfolding
3. beta-sheet oligomers
4. amyloid fibrillar aggregates

Question 21

prion def

Answer 21

proteinaceous Infectious particle

-infection involves a change of secondary structure and conformation in the protein
*converts from a alpha-helical to beta-sheet conformation

-PrP^C: protease-resistant protein, cellular = normal
-PrP^SC: protease-resistant protein, scrapie = bad prion
*can’t be broken down/ cant be destroyed by autoclave

Question 22

Kuru

Answer 22

Fore tribe of Papua New Guinea
-prions PrP^SC transmitted by eating dead love ones brain
-kuru stopped dying when cannibalism was stopped

Question 23

Stanley Pruiser

Answer 23

won nobel prize in medicine for discovering prions

-1st case of infectious agent being a protein ( was not a virus or bacteria)

Question 24

Chronic Traumatic encephalopathy (CTE)

Answer 24

butting heads caused Tau misfolding (precipitation of beta-sheets)

-Alzheimer’s requires the Tau neuron

Question 25

p53

Answer 25

tumor suppressor protein that sense damaged DNA sensor and stops them before they grow -initiates apoptosis

Question 26

what happens when there is a mutation in p53?

Answer 26

causes misfolding in p53 which leads to unregulated cell growth = cancer

Question 27

LOOK AT CHAP 6 PT.3 SLIDES FOR STRUCTURES

Answer 27

OKAY

Question 28

Domains

Answer 28

-independent folding regions within a protein often associated with a certain function -25-100 AA residues -connected to each other by loops -bound by weak interactions between side chains -fundamental unit of TERTIARY STRUC *has hydrophobic core -made of folds??? *in multifunctional enzymes each catalytic activity can be on one of several domains

Question 29

Interfaces

Answer 29

btwn domains provide crevices, grooves, and pockets that make good binding or catalytic sites

Question 30

what are the 3 domains of a polypeptide chain?

Answer 30

1. regulatory domain 2. substrate binding domain 3. nucleotide binding domain

Question 31

4 Protein Classes

Answer 31

1. alpha 2. beta 3. alpha + beta 4. alpha/beta: consists of 2 alpha+beta barrel

Question 32

folds

Answer 32

-combination of secondary structure that form the core of a domain -made of motifs (supersecondary structure)

Question 33

EF-hand fold

Answer 33

-Structure: alpha helix; right hand like an L with fingers pointing towards you *E helix = pointer finger *F helix = thumb -found in proteins that bind Ca^2+

Question 34

Calmodulin

Answer 34

*know structure - has 4 EF-Hand folds

Question 35

Zinc Finger

Answer 35

-made with one alpha and one beta strand -Zinc^2+ in the middle -Zinc is bonded to cysteine and histidine

Question 36

Jelly/ Swiss-Barrel Roll Fold

Answer 36

squished antiparallel beta barrel

Question 37

TIM fold barrel

Answer 37

- alpha/beta barrel -chicken muscle triose phosphate isomerase

Question 38

Rossman Fold

Answer 38

-also known as dinucleotide binding fold -alpha/beta barrel -can bind NAD+ and ATP

Question 39

Horseshoe Fold

Answer 39

-alpha/beta -looks life a horseshoe -Ribonuclease inhibitor

Question 40

Quaternary Structure

Question 41

rotational symmetry

Answer 41

-multisubunit protein display symmetry -defined as Cn *C: cyclical *n: number of subunits -higher order types: octahedral or tethrehedral etc..

Question 42

Dihedral symmetry N-fold

Answer 42

it intersects a two-fold rotational symmetry at right angles