Proteomics Flashcards
What are the goals are proteomics?
To identify and determine the level of proteins, their localization, dynamics, interaction partners and post-translational modifications in specific biological conditions.
What are the levels at which proteins are influenced at?
Think central dogma!
- transcriptional control
- RNA processing control
- RNA transport and localisation control
- Translation control
- mRNA degradation control
- Protein activity control
List some challenges of proteomics
Diverse chemical composition of proteins Large complex structures Changes of abundance over time Different localizations Post-translational modifications Interaction partners
Explain the principles of SILAC
Stable Isotope Labelling with Amino acids in Cell Culture
As part of quantitative proteomics, SILAC provides in vivo labelling of proteins for proteomics studies using mass spec. It involves growing 2 population of cells, one grown in normal culture medium and other in stable heavy isotopes, which results in its uptake into the cells. The heavy isotope will result in heavy aa with higher molecular mass than the control aa, and it will show up as different peaks in the mass spec. This is important in the study of cell-signalling, protein interactions, post-translational modifications, and regulation of gene expressions.
Explain the principles of iTRAQ
isobaric Tags for Relative and Absolute Quantification
Isotope-labelled molecules are covalently-bonded to N-terminus and side-chain amines of proteins after digestion. Each peptide fragment is labelled with a different iTRAQ reagent and combined into a single mixture. The mixture is analysed by MS for identification and quantification. The intensity of the fragments is used to quantify the individual representative peptides
Briefly explain “Label Free” quantitative proteomics
Signal intensities are used for quantification, relies on linear correlation between peak areas of measured peptides and their abundance
