Development and application of advanced proteomic techniques for high-throughput identification of proteins

Abstract

This study aimed to develop and apply advanced proteomic techniques from three aspects for high-throughput identification of proteins, especially enzymes and their associated proteins in yeast proteome (catalomics). Firstly, to validate the high-throughput capacity of DIGE, yeast proteome upon exposure to fifteen kinds of metal stimuli has been interrogated in a parallel and quantitative fashion (quantitative proteomics). Yeast proteins with significantly altered expression levels have been identified, leading to a better understanding of multiple cellular defense mechanisms in yeast. Secondly, to improve the quality of protein-protein interaction data, a new strategy has been developed, whereby subtraction of those nonspecifically bound proteins with the aid of DIGE led to a rigorous identification of yeast metacaspase-binding proteins from yeast proteome (interactomics). Thirdly, to expedite the activity-based protein identification, expression display and phage display have been exploited in attempts to identify the proteins with particular enzymatic activity in a high-throughput manner (functional proteomics).