Chpt 2 continued Flashcards
What allows proteins to fold
Change in conformation (rotation) in the N-Calpha and Calpha-C single bonds in the protein back bone
Phi
is angle of rotation of N-Calpha -80 degrees
Psi
is angle of rotation of Calpha-C +85 degrees
Ramachandran diagram
-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
What amino acids form disulfide bridges/bonds
Cysteine
2 cysteines combine to form
cystine -connected by disulfide bridges
What are the four levels of Protein Structure
Primary, Secondary, Tertiary, Quaternary
Primary Structure
-linear sequence of amino acids
Secondary Structure
Alpha helix Beta Sheets Beta Turns Omega Loop -hydrogen bonding between the carboxyl oxygen and nitrogen hydrogen of the peptide chain (back bone)
Tertiary Structure
-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
Quaternary Structure
interaction of different polypeptide chains (subunits) to form a functional protein -hemoglobin-2 alpha subunits and 2 beta subunits
Alpha Helix
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
Alpha Helix is disrupted by:
- 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)
Proteins that contain alpha helixes
Ferritin -An iron storage protein Hemoglobin -Oxygen caring protein
Beta Pleated Sheets
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
Proteins containing beta sheets
-A fatty acid binding protein - Green fluorescent protein
Loops and Turns
connect secondary structures to form domains -Beta Turn (hairpin or reverse turn) -Omega Loops (or just loops)
Beta Turn
(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
Omega Loops
(or just loops) -No regular repeating structure, but are rigid and well defined -usually on the surface of proteins
Denaturing proteins
unfolding and disorganizing of a proteins secondary and tertiary structure -does not involve hydrolysis of peptide bonds
What are the denaturing agents and reducing agents for Unfolding proteins
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
Amino Acid Modifications -Def
-covalently modified -usually a post translational modification
Amino Acid Modification Types
1) Hydroxylation- add OH 2) Carboxylation- add COO- 3) Glycosylation- add carbohydrates 4) Attach Fatty acids 5) Phosphorylation- add (PO32-)
Alpha Keratin
-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
Collagen
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
Proline in collagen
-facilitates formation of helix by introducing “kinks” in chain -100% trans orientation -Proline and Lysine are often hydroxylated 1) hydroxyproline (Hyp) 2) hydroxylysine (Hly)
Hydroxylation of Proline
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
Hydroxylation of Lysine
Hydroxylysine (Hly) -lysyl hydroxylase (enzyme used) -requires vit C (ascorbate) and molecular oxygen allows cross linking of collagen
Glycosylation of Hydroxylysine
-occurs in some of hydroxylysine residues -attaches carbohydrate to protein adds glucose of glucosylgalactose -glc= glucose -gal=galactose
Triple Helix structure of Collagen
-1000 amino acids in length -3 amino acids per turn -each alpha chain adopts a left handed helix -super helical structure
Biosynthesis in Collagen
Synthesized in: -fibroblasts -osteoblasts of bone -chondroblasts of cartilage
Where does Collagen function?
secreted into the extracellular matrix because it functions outside the cell
Pathway of biosynthesis of collagen
1) rER 2) Golgi Apparatus 3)Secreted outside the cell 4) Collage fibers form (insoluble)
Pathway of biosynthesis of Collagen-rER
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)
Pathway of biosynthesis of collagen- Golgi Apparatus
-Transport in vesicles to plasma membrane
Pathway of biosynthesis of collagen- Extracellular Matrix
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
Types of cross linking in collagen
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)
Degradation of Collagen
-Highly Stable–> half life several months -Remodels due to growth and injury -breakdoown by collagnases (metalloproteinases)
Collagen Diseases
1)Ehlers-Danios Syndrome (EDS) 2) Scurvy ** 3) osteogenesis imperfecta
Ehlers-Danios Syndrome (EDS)
- 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
Scurvy
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
Osteoimperfecta
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
Protein folding
-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
Protein Folding: Denaturation/renaturation experiment
Ribonuclease -124 amino acids -4 disulfide bonds 105 possible combinations- only 1 combination produces 1 enzymatic activity
How does a protein make the transition from unfolded to native form?
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
Protein Folding Diseases
1) Alzheimer Disease 2) Mutant Tau Protein 3) Transmissable Spongiform encephalopathies
Alzheimer Disease
An amyloidose
Amyloid plaque
- involves AB, a 40-43 amino acid peptide
- accumulates in nonbranching fibrils with beta sheet conformation
- neurotoxic
Mutant Tau Protein
-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
Transmissible Spongiform Encephalopathies
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
Names for various TSEs
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)
