Absolute quantitation of protein contents based on the protein expression level indicators converted exponentially by mass spectrometry

摘要

The present inventors have established protein expression index (PAI, π) and in order to determine the protein content of the protein mixture solution using a nano LC-MSMS data. Results digested peptides were analyzed by nano-LCMS / MS, obtained was applied to a Mascot (Mascot) protein identification algorithm based on the tandem mass spectrum. PAI is defined to be a one obtained by dividing the number of peptides found in the number of peptides that can be detected per protein. PAI from serum albumin of different concentrations showed a linear relationship to the logarithm of the protein concentration. This was also valid for 47 proteins of mouse whole cell lysates were analyzed by nano-LC-MS / MS a single time. On the other hand, the number of peptides that have been identified per protein and Mascot protein score, correlation to the protein expression level was bad. For absolute quantitation, PAI was converted (EMPAI, mπ) to PAI that has been modified exponentially proportional to the protein content in the protein mixture. About 47 proteins of the total lysate, the percentage deviation of the concentration of EMPAI base for the value was 63% less than the average actual. EMPAI is successfully applied to a comprehensive protein expression analysis was carried out a comparative study between the gene and protein expression in HCT116 human cancer cells. The present invention provides a method and computer program for quantifying protein content, based on the protein expression level indicator.