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73 Cards in this Set

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What are amino acids (in terms of functional groups)?
-an amino derivative of carboxylic acids (both are linked to the alpha carbon)
What are Zeitterions?
-at physiological pH, the amino group os protonated and the carboxyl is ionized
What are the ploar uncharge amino acids?
-glycine
-serine
-asparagine
-threonine
-cysteine
-glutamine
-tyrosine
What are thh hydrophobic amino acids?
-leucine
-proline
-alanine
-valine
-methionine
-tryptophan
-phenylalanine
-isoleucine
What are the basic amino acids?
-lysine
-histidine
-arginine
What are the acidic amino acids?
-aspartic acid
-glutamic acid
How many of the 20 AA are chiral?
-19 out of 20!
(exist as stereoisomers)
What are stereoisomers?
-they have same molecular formula but different arrangements in space
There are about 200 naturally occurring AA. How can there be so many?
-exist as L-AA's in biochemical pathways
-D-AA's in microbe cell walls and antibiotics
-Post-translationally modified AA
What is the primary sequence of proteins?
-all peptide bonds are the same
-only sequence of side chains is different
-vary widely in size and complexity
Do cells contain lots of protein?
Yes, Rat liver has a lot. E-coli is a close second. Even spinach has protein!
What are the essential amino acids?
-arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tyrptophan, valine
What are the nonessential amino acids?
-alanine, asparagine, aspartate, cysteine, glutamate, glutamine, glycine, proline, serine, tyrosine
Which two essential amino acids are actually only needed in juveniles of rats and humans?
-arginine
-histidine
How does a Ramachandran plot work?
-only white spots are possible bond angles.
What is amino acid analysis?
-finding what amino acids are present and in what amounts in a protein
What is the sequence of amino acids (not analysis)
-finding the unique sequence of amino acids in a protein
How do you perform acid hydrolysis of peptides?
1) make up in 6 N HCL
2) heat at 110 C for 24 hours
What are different ways of separating out amino acids?
-peptide hydrolysis
-chromatography
-chromatogram
-the Ninhydrin reaction
Is the peptide bond flat or bent? Why?
-bond lies flat due to double bond character (DBC).
*Double bond moves between N=C and C=O
For the 20 amino acids we learned, what conformation are they even though they're not drawn that way?
-'L' acids
What are two amino acids that are rarely found in proteins?
-tryptophan
-thyamine
Which amino acid usually has disulfide bonds?
-cystein
what two groups will be destroyed if they are put in an acid hydrolysis?
-glutamine
-asparage
(amide groups get hydrolized)
*tryptophan can't stand it either
What is the point of Ninhydrin in a chromatogram?
everything turns purple except proline which is yellow
What does the height indicate of the peaks in a chromatogram?
-amount of amino acid present
What are 4 chromatographic methods?
1) adsorption
2) ion exchange
3) partition
4) gel permeation
How does ion exchange work?
-AA's interact with a stationary charge
How does partition work?
-use of a liquid that separates objects
What does gel permeation do?
-acts as a molecular sieve by separating large sizes first
What is the problem with gel permeation?
-it's hard to find a medium that won't cause adsoption or ion exchange
Once again (and then some), what are the problems with acid hydrolysis?
-tryptophan destroyed
-glutamine and asparagine hydrolysed
-serine and thronine lowered (desaturation)
Why do scientists still use this method?
-testing facilities insist on it!
How is the procedure for Ion exchange chromatography?
1)add cation exchange bead before sample
2) add mixture of Asp, Ser, Lys
3) add Na+ (such as NaCl): Asp lets go first due to least positive charge
4) increase Na+: Ser elutes next
5) increase Na+ more: Lys will elute last since it has highest charge
What protein is sometimes able to denature and renature?
-RNAase
What are two major protein sequencing reagents?
-Sangers Reagent (DNFB)
-Dansylchloride
What are 4 major protein cleaving reagents?
-Chymotrypsin
-Trypsin
-Endoprotease V8
-Cyanogenbromide
Where do cleaving reagents cleave?
-on carboxy side of group
How does Chymotrypsin work?
-cleaves on carboxy side of: Phe, Tyr, and Trp
-works on the aromatics
How does Trypsin work?
-cleaves on carboxy side of: Lys and Arg.
-works on basic groups
How does Endoprotease V8 work?
-cleaves on carboxy side of: Glu or Asp
-works on strong acidic groups
How does Cyanogenbromide work?
What is different with this reagant from the others?
Converts Met into homoserine
*Cyanogenbromide is a chemical reagant NOT an enzyme, nor does it cleave.
What is the major difference between a twisted fiber and a pleated fiber?
-twisted can stretch to twice the length and back again; pleats barely increase before returning to form
Where do the characteristics of fibrous proteins come from?
-a result of their sequence and repeated bond angles
What amino acids make up silk fibroin and what is their ratio?
-gly, ala, ser
-3:2:1
ex; g-s-g-a-g-a-g-s-g-a-g-a-g-s
What shape is silk fibroin?
-found in beta pleated sheets
How does silk stretch in terms of wool?
-very poorly
-doesn't bounce back
What makes up alpha keratins?
-alpha helices (this is how you would make string into a rope)
What component of wool makes it so stretchy?
-sulfur content: it has cystein which has disulfide bonds
Is wool an alpha keratin?
Yes
What item makes up 25-50% of the protein in vertebrates?
-collagen
What are the key characteristics to collagen?
-very insoluble
-strongly resists stretching
-has lots of gly and pro (or hydroxypro which is more stable)
What is the sequence for collagen?
-mostly repeating Gly-X-Pro
Why is there so much Gly found in collagen?
-it is woven tightly and only Gly can fit in the center where the twists meet
What are the fours levels of protein stucture?
-primary
-secondary
-tertiary
-quartenary
How many levels of protein structure does hemoglobin show?
-all four levels
Describe the first level of protein structure: primary
-primary structure is the order of AA's in the polypeptide
Describe secondary structure.
-secondary structure is regularities in local conformations
Describe tertiary structure.
-tertiary structure is the completely folded protein, interactions may be between distant parts of the sequence
Describe quarternary structure.
Quarternary structure subunit interactions in an oligomeric protein
What causes Sickle-Cell anemia?
-An alteration in the sequence of the hemoglobin beta-chain at position 6 from Glu to Val
What affect does the change in hemoglobin have on shape of RBC's?
Deoxyhemoglobin is more hydrophobic in one area and this leads to the formation of long rigid fibers
What is denaturation?
-disruption of native conformation of a protein, with loss of biological activity
-can be caused by heating or chemicals (very small energy)
How can denaturation be monitored?
-monitor changes in ultraviolet (blue), viscosity (red), optical rotation (green).
How does 'in vivo' (inside cell) protein folding occur?
-folded proteins occupy a very low-energy well which makes it very stable
-proteins fold spontaneously into this conformation
-folding continued by previous folds through interactions
-folding is extremely rapid <1s
-
As a protein is folding, what happens to its shape and why?
-the polypeptide collapses in upon itself due to the hydrophobic effect
-an intermediate "molten globule" forms with elements fo secondary structure
-backbone rearranged to acheive a stable native conformation
What is RNase?
-Anfinsen denatured it using 8 M urea and beta mercaptoethanol (ME)
-then he removed them to allow refolding (it didn't work)
-he added catalytic amounts of beta ME and RNase reformed to 100% of its activity and function! He won the nobel.
What does the hydrophobic effect contribute in detail to protein folding?
-nonpolar side chains associate causing a polypeptide chain to collapse to the molten globule
-nonpolar chains inside while polar and charged side chains remain on surface facing water
What is the driving force for protein folding?
-the LARGE increase in entropy from water released to bulk solvent!
*hydrophobic collapse of a polypeptide occurs at the same time as formation of secondary structures
How does hydrogen bonding contribute to protein folding?
-contributes to cooperativity of folding
-helps stabilize secondary structures and native conformation
How do Van der Waals interactions contribute to protein folding?
-VDW contacts occur between nonpolar side chains and contribute to stability
-Charge-Charge interactions between oppositely charged side chains in protein interior also may stabalize
What is Anfinson's Flicker hypothesis?
I don't know!
-prepared antiserum to native Staph Nuclease or to a peptide fragment of aminoacids I-99
How did Anfinson go about his Flicker Hypothesis?
-purified antibodies to peptides through affinity chromatography
-Introduced antibodies into native conformation over the peptide column
-had same effect on native form as it did from the peptide