Chpt 2 continued

Question 1

What allows proteins to fold

Answer 1

Change in conformation (rotation) in the N-Calpha and Calpha-C single bonds in the protein back bone

Question 2

Phi

Answer 2

is angle of rotation of N-Calpha -80 degrees

Question 3

Psi

Answer 3

is angle of rotation of Calpha-C +85 degrees

Question 4

Ramachandran diagram

Answer 4

-displays favored and disfavored phi and psi bond angle combinations -many conformations are disallowed due to steric hindrance (steric exclusion) -Left handed helixes are rare

Question 5

What amino acids form disulfide bridges/bonds

Answer 5

Cysteine

Question 6

2 cysteines combine to form

Answer 6

cystine -connected by disulfide bridges

Question 7

What are the four levels of Protein Structure

Answer 7

Primary, Secondary, Tertiary, Quaternary

Question 8

Primary Structure

Answer 8

-linear sequence of amino acids

Question 9

Secondary Structure

Answer 9

Alpha helix Beta Sheets Beta Turns Omega Loop -hydrogen bonding between the carboxyl oxygen and nitrogen hydrogen of the peptide chain (back bone)

Question 10

Tertiary Structure

Answer 10

-Folding of polypeptide chain as a result of interactions between R-groups: these interactions can be-> disulfide bond, hydrophobic interactions, Hydrogen bonding, and ionic bonding -A domain is a unit of tertiary structure 1) helix turn helix 2) helix loop helix 3) zinc fingers 4) leucine zipper

Question 11

Quaternary Structure

Answer 11

interaction of different polypeptide chains (subunits) to form a functional protein -hemoglobin-2 alpha subunits and 2 beta subunits

Question 12

Alpha Helix

Answer 12

Secondary Structure -Orientation is right handed helix -stabilized by hydrogen bonding (every fourth amino acid) between carbonyl carbon and NH group of peptide -3.6 amino acids per turn of helix -R- group extended outward -Helix is disrupted by: 1) proline 2) large number of charged amino acids (Q, E, H, K, R) 3) Amino acids with bulky side chains (W) 4) Amino acids with branched R groups

Question 13

Alpha Helix is disrupted by:

Answer 13

1. Proline 2. Large number of charged amino acids (Q, E, H, K, R) 3. Amino acids with bulky side chains (W) 4. Amino acids with branched R groups (V, I)

Question 14

Proteins that contain alpha helixes

Answer 14

Ferritin -An iron storage protein Hemoglobin -Oxygen caring protein

Question 15

Beta Pleated Sheets

Answer 15

Secondary Structure -orientation of proteins is flat or pleated, linear “sheet” of proteins -stabilized by hydrogen bonding between carbonyl oxygen and NH group of peptide -Adjacent amino acids are separated by 3.5 A -Strands may organize themselves into several orientations: Antiparallel, Parallel, Mixed -Beta bends: contain proline and glycine

Question 16

Proteins containing beta sheets

Answer 16

-A fatty acid binding protein - Green fluorescent protein

Question 17

Loops and Turns

Answer 17

connect secondary structures to form domains -Beta Turn (hairpin or reverse turn) -Omega Loops (or just loops)

Question 18

Beta Turn

Answer 18

(hairpin or reverse turn)–> connect secondary Structures to form domains: -stabilized by h-bonding between Co group of residue 1 and NH group of residue three amino acids down -usually on the surface of proteins

Question 19

Omega Loops

Answer 19

(or just loops) -No regular repeating structure, but are rigid and well defined -usually on the surface of proteins

Question 20

Denaturing proteins

Answer 20

unfolding and disorganizing of a proteins secondary and tertiary structure -does not involve hydrolysis of peptide bonds

Question 21

What are the denaturing agents and reducing agents for Unfolding proteins

Answer 21

Denaturing Agents: -heat -organic solvents -urea -guanidium chloride -detergents–> SDS -change in pH–> Strong acids and Strong Bases -Heavy Metals–> Pb or Hg Reducing Agents -Beta mercaptoethanol->reduces disulfide bonds

Question 22

Amino Acid Modifications -Def

Answer 22

-covalently modified -usually a post translational modification

Question 23

Amino Acid Modification Types

Answer 23

1) Hydroxylation- add OH 2) Carboxylation- add COO- 3) Glycosylation- add carbohydrates 4) Attach Fatty acids 5) Phosphorylation- add (PO32-)

Question 24

Alpha Keratin

Answer 24

-Primary component of hair, wool, horns, claws, and hooves -Two right handed alpha helixes entwined to form a coiled:coiled structure. Wrapped around each other in a left handed manner -Helixes are cross linked by: 1) Van Der Waals forces 2) Ionic Interactions 3) Disulfide bonds -Contains heptid repeat–>each alpha helix contains 3.5 residues per turn -Hair and wool would have fewer disulfide bridges–>makes them stretchable -Horns, claws, and hooves have more disulfide bridges–> makes them harder

Question 25

Collagen

Answer 25

A fibrous Protein -most abundant protein in the body -composed of long rigid ALPHA chains (not alpha helix) wrapped around each other in a left handed triple helix (rope like) -Repeating Triplet–> Glycine, Proline, X- where x Is any amino acid

Question 26

Proline in collagen

Answer 26

-facilitates formation of helix by introducing “kinks” in chain -100% trans orientation -Proline and Lysine are often hydroxylated 1) hydroxyproline (Hyp) 2) hydroxylysine (Hly)

Question 27

Hydroxylation of Proline

Answer 27

Hydroxyproline (Hyp) -Prolyl hydroxylase (enzyme used) -requires vit C (ascorbate) and molecular oxygen location of hydroxylation -4-hydroxyproline (more) -3-hydroxyproline (less) allows cross linking of collagen

Question 28

Hydroxylation of Lysine

Answer 28

Hydroxylysine (Hly) -lysyl hydroxylase (enzyme used) -requires vit C (ascorbate) and molecular oxygen allows cross linking of collagen

Question 29

Glycosylation of Hydroxylysine

Answer 29

-occurs in some of hydroxylysine residues -attaches carbohydrate to protein adds glucose of glucosylgalactose -glc= glucose -gal=galactose

Question 30

Triple Helix structure of Collagen

Answer 30

-1000 amino acids in length -3 amino acids per turn -each alpha chain adopts a left handed helix -super helical structure

Question 31

Biosynthesis in Collagen

Answer 31

Synthesized in: -fibroblasts -osteoblasts of bone -chondroblasts of cartilage

Question 32

Where does Collagen function?

Answer 32

secreted into the extracellular matrix because it functions outside the cell

Question 33

Pathway of biosynthesis of collagen

Answer 33

1) rER 2) Golgi Apparatus 3)Secreted outside the cell 4) Collage fibers form (insoluble)

Question 34

Pathway of biosynthesis of Collagen-rER

Answer 34

1) translation (protein synthesis) on bound ribosomes -begins life as free ribosome in cytosol -cleavable, N-terminal signal sequence–>prepro alpha chain
2) Post translational modification
- signal peptidase removes signal sequence to form pro alpha chain
- hydroxylation of proline or lysine in “y” position->Gly-x-y -glycosylation of (some) hydroxylysine residues-> glucose and glucosyl-galactose
- Procollagen is formed by “wrapping” together of pro alpha chains-> formation of interchain disulfide bridges in C-terminal extensions by protein disulfide isomerase
- Transports to the Golgi by vesicles in the secretory pathway (bulk flow)

Question 35

Pathway of biosynthesis of collagen- Golgi Apparatus

Answer 35

-Transport in vesicles to plasma membrane

Question 36

Pathway of biosynthesis of collagen- Extracellular Matrix

Answer 36

Secretion outside cell (extracellular matrix) -exocytosis -cleavage of C and N terminal precollagen peptides by N and C pro collagen peptidases and mature into mature collagen “tropocollagen” -crosslinking

Question 37

Types of cross linking in collagen

Answer 37

1)deamination of lysine and hydroxylisine -lysyl oxidase, produces aldehyde derived intermolecular crosslinks–>allysine and hydroxyallsine residues 2) aldol condensation products -aldehyde + alehyde 3) Schiff Base (imine) cross links -aldehyde + amine (R group of lysine)

Question 38

Degradation of Collagen

Answer 38

-Highly Stable–> half life several months -Remodels due to growth and injury -breakdoown by collagnases (metalloproteinases)

Question 39

Collagen Diseases

Answer 39

1)Ehlers-Danios Syndrome (EDS) 2) Scurvy ** 3) osteogenesis imperfecta

Question 40

Ehlers-Danios Syndrome (EDS)

Answer 40

• heterogenous group of generalized connective tissue disorders (about 10)
• Heritable defects in fibrillar collagen:
  1) deficiency of collagen processing enzymes
  2) mutations in amino acid sequence of collagen 1, III, or V. A)Mutation of type III collagen= normal component of arteries, degraded or accumulates in cells, potentially lethal vascular problems

B)mutation of type I collagen= normal component of skin and joints, stretchy skin and joints

Question 41

Scurvy

Answer 41

the reason british sailers are called lymes!

1) Reduce tensile strength of collagen

A)defficiency of ascrobic acid (vit C) inhibits Prolyl Hydroxylase and Lysyl hydroxylase

B) collagen lacks cross links

2) symptoms

bleeding (leaky) capillaries

Question 42

Osteoimperfecta

Answer 42

1-30,000 to 50,000 are born with

• brittle bone disease
• due to defects in synthesis of collagen I–>mutation of alpha 1 or alpha 2-substitution of amino acid other than glycine which prevents proper folding of alpha chain into helix (bulky side chains)
• Symptoms: fragile bone, thin skin, abnormal teeth, weak tendons

TWO TYPES

1) osteogenesis imperfecta tard–>type I collagen defect; presents in early infancy, fractures secondary to minor trauma
2) osteogenesis imperfecta congenita–> type I collagen defect; patients die in utero or neonatal period due to pulmonary hypoplasia

Question 43

Protein folding

Answer 43

-all the information necessary for a protein to fold into its final 3-D structure is contained in the amino acid sequence -different secondary structures contain diff quantities of amino acids -Bring amino acid R-groups together at the active site. The R group at the active site can be from distant region of primary sequence -studies suggest “all or nothing process” due to cooperative transition. A rapid transition from folded (native) to unfolded (denatured) state

Question 44

Protein Folding: Denaturation/renaturation experiment

Answer 44

Ribonuclease -124 amino acids -4 disulfide bonds 105 possible combinations- only 1 combination produces 1 enzymatic activity

Question 45

How does a protein make the transition from unfolded to native form?

Answer 45

1) Random Search Model -100 residue proteins would take 1.6x10^27 years to fold 2)Cumulative selective Model -the essence of protein folding is the tendency to retain partly correct intermediates

Question 46

Protein Folding Diseases

Answer 46

1) Alzheimer Disease 2) Mutant Tau Protein 3) Transmissable Spongiform encephalopathies

Question 47

Alzheimer Disease

Answer 47

An amyloidose

Amyloid plaque

• involves AB, a 40-43 amino acid peptide
• accumulates in nonbranching fibrils with beta sheet conformation
• neurotoxic

Question 48

Mutant Tau Protein

Answer 48

-Tau is the protein produced from translation of an alternatively spliced gene called MAPT (microtuble-associated protein tau) located in humans on chromosome number 17

Question 49

Transmissible Spongiform Encephalopathies

Answer 49

Misfolded Prion Proteins -normal prion found in neurons and glial cells -lack primary sequence differences or post translational modification differences from normal prion protein -apparently the altered form becomes the template to induce misfolding of normal PrP into altered form

Question 50

Names for various TSEs

Answer 50

Transmissible Spongiform Encephalopathies Human -Creutz-Jakab disease classic or vCJV -Kuru Cattle -Bovine spongiform encphelopathy (BSE or mad cow) Sheep -scrapie Deer family -chronic wasting disease (CWD)