chapter 3 Flashcards
the Merrifield method
one end of peptide = attached to resin in column
protective chemical groups block unwanted reactions
Chemical synthesis of peptides by the Merrifield method:
proceeds from the carboxyl terminus to the amino
terminus.
amino acid sequence can inform:
3D structure
function
cellular location
evolution
consensus sequence =
reflects most common amino acid at each position
bioinformatics:
identifies functional segments in new proteins
establishes sequence and structural relationships to known proteins
horizontal gene transfer
= transfer of a gene or group of genes from one organism to another
proteins derived from transferred genes are not good candidates for bacterial evolution studies
for example, rapid spread of antibiotic-resistance genes in bacterial populations
Homologs from different species are called
Orthologs are homologs found in the same
signature sequences
certain protein segments specific to a taxonomic group
The amino acid sequences of proteins:
can be used to establish evolutionary relationships
between organisms.
Proteins Can Be separated based on:
size
charge
binding properties
protein solubility
purifying proteins steps
- crude extract
- fractionaction
- dialysis
electrophoresis can estimate
- number of proteins in a mixture
- degree of purity
-isoelectric pt - molecular weight
proteins migrate based on
charge to mass ratio
isoelectric focusing
A protein sample is added on one end of a strip with a pH gradient. A charge is added and proteins migrate across the strip, stopping at their isoelectric point.
2D electrophoresis
- proteins are first separated by isoelectric focusing
- the gel strip used for the isoelectric focusing is laid horizontally on a second slab gel.
- proteins are separated vis SDS electrophoresis
the horizontal phase separates by pI
and vertical separates by molecular weight (SDS)
protein spots can be cut out and identified via mass spectrometry
electrophoresis without SDS separates based on what
a function of size and shape
electrophoresis with SDS separates based on what
molecular weight. SDS binds to and unfolds proteins, making charge to mass ratio approx equal in the proteins. They are then able to be separated by molecular weight.
smaller proteins mitigate faster
electrophoretic mobility equations
velocity/ potential. (V/E)
Net charge/ frictional coeffecient (Z/T)
what will increase during purification of an enzyme
specific activity.
non polar amino acids
GAPVILM
negative amino acids
Asp, Glu
positive amino acids
His, Arg, Lys
conjugated proteins
contain permanantley associated chemical compounds
_______ bonds link amino acids in a polypeptide chain while __________ bonds link the polypeptide chains
covalent, non-covalent
