export_aa Flashcards
Hydrophobic amino acids
* GAV LIMP TP * Gly, ala, val, leu, I le, pro, phe, met, trp
Polar amino acids
* Polar CATTS G rowl * Cysteine, asparagine, threonine, tyrosine, serine, glutamine
Polar uncharged amino acids
* Thre e TAGS * Threonine, tyrosine, asparagine, glutamine, serine
Acidic amino acids
Asp and glu
Basic amino acids
* His lys ar e basic * Arg, his, lys
3 AA that contribute to UV absorbance at λ=280 nm
- Trp (ε ~5000 M -1 cm -1 ) 2. Tyr (ε ~1000 M -1 cm -1 ) 3. Phe (ε ~250 M -1 cm -1 )
Structures at half and full E.P. (equivalence point)
https://s3.amazonaws.com/classconnection/833/flashcards/4068833/gif/half_and_full-153E2F7BA226388E880.gif
Iso (pI) electric point
Characteristic pH where net charge of a protein is 0
To calculate pI of asp/glu
Take an average of sidechain pKa + α-COOH pkA
To calculate pI of basic amino acids
= (side chain pKa + α-NH3)/2
Excluding basic/acidic side chains, for other ionizable groups, how do we calculate pI
- Determine middle pKa 2. Add with α-COOH 3. Divide by 2
pKa range for ammonium (NH3+)
8.8-11.0
pKa range for -COOH
1.8-2.4
Primary protein structure
* Series of aa joined by peptide bonds * Most important aspect of 1° structure is sequence of aa
Secondary protein structure
* Classified (α-helix, β-sheet, β-turn) according to how the polypeptide folds upon itself
Alpha helix
* 3.6 residues per turn * Hydrogen bonding stabilizes the structure (between N-H and O=C)
Beta sheet
* Parallel: β-strands line N-N and C-C terminals * Anti parallel: β-strand folds upon itself, lining N with C * Stabilized by H bonds
Beta turn
* 5 amino-acid residues or less * Lye on the protein surface because they are hydrophilic
Tertiary structure
* Higher order of folding * Stabilized by H-bonds, VDW interactions, disulfide bridge formation (only in cys), and hydrophobic packing
Quaternary structure is described by
* Interaction between tertiary polypeptides * Named according to # of sub-units
What is unique about C-N bond
Has partial double bond character, cannot rotate freely
Rotation is permitted about the
N-Cα and Cα-C bonds
Ramachandran plot
* Determines stable configurations of phi and psi angles * Only a few configurations are energetically favorable, therefore polypeptide chain flexibility is limited
3 characteristic areas on a Ramachandran plot
https://s3.amazonaws.com/classconnection/833/flashcards/4068833/gif/ram-153E35363B36F397A88.gif
Hydrophobic effect
* CH3 (non-polar) does not interact well with H20 * Interaction decreases entropy * When contact is minimized entropy increases due to clustering of non-polar groups
Stabilizing interactions: hydrogen bonding
NH and OH groups form H bonds
Stabilizing interactions: ionic interactions
* In the 2 acidic aa (asp, glu), occurs in non α-COOH * Also occur in a molecule (ie lys) with an extra NH2 group * Ionic bonds form between +&- groups
Stabilizing interactions: disulfide bond
Form only when 2 cysteine residues bond
Gel filtration (size-exclusion) chromatography
Small proteins are trapped first in the beads within the column, while the biggest proteins travel the farthest/fastest
Ion exchange chromatography
* Separation based on charge and the pH of the matrix
SDS PAGE
* SDS binds to protein, giving them all - charge * smaller proteins move fastest * polyacrylamide has bigger pores than agarose
Affinity chromatography
* Separate based on affinity to bind to ligand * Unwanted proteins exit while protein of interest is held in the column
Basic idea of Solid Phase Peptide Synthesis
* since protecting group for C-terminus does not need to be removed, attach C-terminus to resin (solid support or bead) * Add each additional amino acid to free N-terminus of growing peptide chain * Cleave peptide product off solid support as final deprotection
