Methods for Analysis and Quantification of Signal Transduction Proteins in the Yeast Pheromone Pathway
Sponsor: National Institutes of Health
Contact: David Camp
Collaborator: Roger Brent, Molecular Sciences Institute
The overall goal of the Alpha Project at the Center for Genomic Experimentation and Computation is to gain the ability to predict the behavior of a well-studied biological regulatory system at the level of individual cells. PNNL's role in this project is to develop MS-based methods to analyze and quantify phosphorylated and unphosphorylated species of signal transduction proteins in the yeast pheromone pathway. Efforts include:
- quantifying changes in protein kinase activity by changes in phosphorylation of substrates
- identifying new kinase substrates
- generating a comprehensive set of phosphorylated peptides from the yeast proteome
- identifying individual peptides to generate partial sequence data. These experimentally determined peptides are to be compared with a computer-generated database of a theoretical enzymatic digest of all proteins encoded in the yeast genome.
This ability to experimentally measure molecular species from populations of cells provides backup for single cell techniques that do not work in addition to helping calibrate those that do.