proteins Flashcards
Give examples of candidate protein methods
Western analysis
ELISA
immunohistochemistry
Give examples of proteome wide and purification methods
chromatography protein micro-arrays 2D-difference in gel electrophoresis SILAC mass spectrometry
Describe Crick’s central dogma
describe basis of translation
must study proteins - lots of post-translational modifications
not information stored in DNA or RNA
Describe the correlation of mRNA and protein
some well correlated
some not at all
some negatively correlated
What mechanisms cause there to be discrepancies in the quantity of mRNA vs protein?
Control of transcription
miRNAs
Efficiency of mRNA translation - related to ribosomal occupancy
protein decay
Describe a BCA assay
quantifies TOTAL protein
exhibited by a colour change in proportion to the amount of protein
Describe ELISA
Enzyme linked immunosorbent assay
uses colour change and standardised curve
detects and quantifies a specific protein (antigen) in sample
crude method
if commonality in isotope can give inaccurate results
very commonly used
simple
Describe western blots
proteins can be separated by SDS-polyacrylaminde gels, electrophoresis - much higher resolution than mRNA or DNA
separates proteins by size
more than one replicate
use control of stable protein - loading control
Describe immunohistochemistry
e.g. parkin in parkinsons disease
use control - check detecting correct protein
protein localisation in the cell
Describe activated signalling pathways in disease
can monitor phosphorylation of proteins over minutes using Western analysis
This is a dynamic process
may be different in disease strains
What is an easy way to purify proteins?
using genetically-engineered stages
antigenic determinant
GST fusion complex
Give examples of monogenic forms of hypotension and hypertension
BArter syndrome
Liddle syndrome
Gordon’s syndrome
Gitelman’s
How can SNPs be used to inform drug treatment choices?
Proteins involved in the metabolism of drugs etc CYP2CP and CYP2D6 metabolise codeine and warfarin (and a huge range of drugs) mutations in these can have drastic impacts on the metabolism of the drug
What is the goal of proteomics?
to form a comprehensive, quantitative description of protein expression and its changes under the influence of biological perturbations such as disease or drug treatment
What are the major limitations of antibody microarrays compared to DNA and RNA?
too many functional types of proteins
harder to get specific result
different conditions when antibody will react with target protein
