P2.1 Reactions + denaturation
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are proteins more sensitive to their environment than lipids and carbs? + explanation
yes! can undergo drastic changes in structure (denaturation) under relatively mild conditions –> unfolding of tertiary structure and unraveling of secondary structure
is denaturation good or bad for protein?
can be good (ie eggs) and bad (overheat and precipitation) depending on one’s objective
definition of denaturation?
change in structure or conformation of a protein that does not involve breaking of peptide bonds (primary structure remains intact)
secondary and tertiary structure involves a balance of a number of ____________ bonds/interactions (4 types?) that hold protein in a space - most interactions are ______ but their high numbers provide _________ - denaturation results from the __________ of these interactions
• non-covalent • hydrophobic interactions, electrostatic interactions, hydrogen bonds, van der waals forces • weak, provide stability • disruption
sensitivity of proteins to denaturation is a function of many variables: (5) - all also govern what of protein?
• amino acid composition • chain length • complexity of its structure • prosthetic groups • environmental conditions (ie pH) all govern functionality of protein
casein has little _________ structure –> consequence? vs egg albumin
• little secondary structure • does not denature readily even when boiled, vs globular egg albumin is very sensitive to temperature
when the conformation of a protein is changed, many of its ________ properties are also changed
physical
5 common changes observed when denaturation takes place
1) unfolding of the tertiary structure/unraveling of the secondary structure –> makes the peptide bonds more accessible to proteolytic enzymes 2) denaturation of protein generally results in a reduction in its solubility (bc hydrophobic core is out) –> can precipitate and form a gel 3) enzymatic activity may be decreased or lost 4) viscosity generally increases, gelation may occur 5) crystallization is no longer possible (no 2°, 3° or 4° structure)
if denaturation is extensive, what happens? (3)
• globular form can be completely unraveled and its secondary structure badly disrupted • many reactive groups like COOH, NH2, SH, etc. are exposed –> capable of reacting with each other, so that polymerization or aggregation takes place • some proteins form gels when denatured, while others precipitate out of solution
difference between aggregation and polymerization?
• aggregation: protein coprecipitate with each other –> hydrophobic and sulfite bond • polymerization: carboxyl and amino group attach from 2 different proteins –> need high temp and catalyst
most common factor contributing to denaturation? explanation
heat! - increase in kinetic energy of molecules result in disruption of H-bonds, which are relatively weak
other important factors that contribute to denaturation (besides heat and more general) (4 + 1)
• changes in pH • changes in ionic strength • chemical agents (ethanol, acetone, urea) • surface forces (ie. add air to proteins) • combinations of above
can the disruption of 1 alpha helix change 3° structure?
yes
most proteins tend to be stable over a relatively large/narrow pH range
narrow
shift in pH affects ________ ________ of molecule, which affects the __________ bond contributions to the tertiary structure
• overall charge • electrostatic bond
proteins with significant quantities of (3 aa) are especially susceptible to changes in pH
• aspartic acid • glutamic acid • lysine
many soluble proteins _________ when they are subjected to environmental conditions close to their _________ point - why?
• precipitate • isoelectric • proteins fall out of solution because they don’t interact with water anymore –> positive and negative charges are balanced
contributors to denaturation (5)
1) effect of pH 2) Hydrogen bond breakers 3) Detergents 4) Organic solvents 5) surface forces
compounds that can effectively compete with _______ linkage for H-bonds can readily disrupt both ______ and _______ structure - ex.: ?
• peptide linkage for H-bond • both 2° and 3° structure • ex.: urea
urea is commonly used to _______ proteins for analysis by _________, which is based on the fact that migration of proteins in an _______ _______ depends on their ____ and ___ ________ –> if not _________, their shape would be a confounding factor affecting their migration in the _______ ________
• denature • by electrophoresis • electric field • depends on their size and net charge • not denatured • the electric field
what are good H-bond disruptors? how? mostly used for what?
alcohol and acetone - by partial dehydration of a protein - however, the degree of structure modification is not as severe as urea - mostly used to precipitate enzymes from solution for isolation with minimal denaturation
detergents have both (2) moieties - can also be called a ? - ex? used in what?
• hydrophobic and hydrophilic moieties • a surfactant • ex.: sodium dodecyl sulfate (SDS) –> used in electrophoresis
detergents are able to bridge the (2) regions of protein = ________ of the structure of the protein
• hydrophobic and hydrophilic • opening
organic solvent can make the environment hydrophobic/hydrophilic –> what does that do to the protein? (2) - ex of organic solvent?
• hydrophobic • can turn the protein inside out • can also make enzymes work in totally opposite ways –> make lipase add FFAs to glycerol • hexane