Proteins and Peptides

peptides

short polymers of amino acids (less than 50 amino acids long)

5 kDa

average molecular weight of a polypeptide

amide bonds

form between carboxylic acids and amines; polar and neutral

peptide bond

type of amide bond that forms link between terminal amine of one amino acid and the carboxylic acid of another

tautomerize

peptide bonds ________________ giving them double bond character that makes them have restricted peptide confirmations

proteins

polymers of amino acids

53 kDa

proteins are about 10x larger than peptides with an average molecular weight of __________ (about 480 amino acids per molecule)

primary structure

sequence of amino acids from N- to C- terminus

secondary structure

folding of short (3-30) amino acids into geometrically ordered units

alpha helix

type of secondary structure that occurs when hydrogen bonds form between the hydrogen of an amine and the carbonyl oxygen of another amino acid with the residues pointing outwards

beta sheet

type of secondary structure in which the amino acids are aligned in a zig-zag; peptides are flat

tertiary structure

secondary structures assemble into larger functional units such as the mature polypeptide and component domains; form hydrogen bonds, ionic bonds, disulfide bridges, and hydrophobic interactions

quaternary structure

clustering of two or more individual polypeptides

protein maturation

process of reaching tertiary and quaternary confirmations

post-translational modification

addition of new chemical groups or removal of transiently needed peptide segments and stabilizing interactions in the maturation process

mature proteins

have function and structural roles such as:

-catalyzing metabolic reactions

-signalling/regulatory molecules

-enabling cellular motion

-provide structure and integrity

-receptors

scurvy

ascorbic acid deficiency resulting in insufficient synthesis of collagen

Menkes disease

disorder in which there is a copper ion deficiency

denaturation

the process in which proteins lose structural integrity without breakage of a peptide bond; compromise hydrogen bonding and disulfide bridges due to heat, pH, organic solvents, urea, heavy metal ions, and detergents

can be reversible in some cases but usually isn't

protein crashing

adding organic solvent to precipitate the protein from solution

domains

fundamental and functional 3D structural units of polypeptides

chaperones

guide protein production and folding in proteostasis (aka heat shock proteins)

protein misfolding

can result in denatured proteins leading to conditions like amyloid diseases or prion diseases

amyloid diseases

Alzheimer's and Parkinson's; result from protein misfolding

prion diseases

bovine spongiform encephalopathy (mad cow), Creutzfeldt-Jakob diseases, Scrapie

stabilizing interactions

involved in protein folding

ex: disulfide bridges, hydrophobic interactions, hydrogen bonds, ionic bonds (salt bridge)

soluble proteins

type of protein that can be extracted from cells using aqueous solutions of physiologic pH and ionic strength

integral membrane proteins

type of protein in which extraction requires dissolution of the cell membrane with detergent because it is a component of the cell structure

globular proteins

type of protein that is compact, roughly spherical molecules that have axial ratios of less than 3; most types of enzymes

fibrous proteins

type of protein that is a structural protein that adopts highly extended conformations; has axial ratio of greater than 10

conjugated proteins

proteins with covalently attached molecules

ex: lipoprotein, glycoprotein, metalloprotein

SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis)

method of identification of complex protein mixtures and determination of their molecular weights

SDS

dissolves and puts negative charge on protein so that they can migrate towards positive electrode in gel

isolation

two main _____________ methods: selective precipitation and chromatographic techniques

selective precipitation

isolation method that exploits differences in relative solubility of individual proteins as functions of pH, polarity, or salt concentration

isoelectric precipitation

type of selective precipitation that separates protein by pH

precipitation with ethanol or acetone

type of selective precipitation that separates protein by polarity

salting out with ammonium sulfate

type of selective precipitation that separates protein by their salt concentration

chromatographic techniques

isolation methods that separate one protein from another using specific columns based on difference in size, charge, hydrophobicity, or ability to bind a specific ligand

size exclusion

type of chromatographic isolation that separates protein by size

ion exchange

type of chromatographic isolation that separates protein by charge

hydrophobic interaction

type of chromatographic isolation that separates protein by hydrophobicity

affinity

type of chromatographic isolation that separates protein by ability to bind to specific ligand

Edmen degredation

reaction that plucks off one amino acid at a time in a polypeptide so it can be identified and sequenced

phenylisothiocyanate

Reactive reagent used in Edman degradation that interacts with the amino group of the first amino acid on the N terminus, breaking the peptide bond and forming a cyclic phenylthiohydantion-(AA)

HPLC

high performance liquid chromatography; used to identify the phenylthiohydantion-(AA) complex (UV active) to identify the amino acid

sanger sequencing

used to determine the primary structure of insulin

mass-spectrometry

gives accurate molecular weight of a protein

NMR

gives protein structure in solution

X-ray crystallography

gives accurate tertiary and quaternary protein structures

proteomics