Response to the Comments on "Determining Allele-Specific Protein Expression (ASPE) Using a Novel Quantitative...

Russell and colleagues deserve credit for being the first to use a QconCAT standard to simultaneously quantify both the wild-type and mutant peptides of a protein (i.e., CYP2B6) ( J. Proteome Res. 2013, 12 (12), 5934-5942. DOI: 10.1021/pr400279u). However, the rationale of their study was entirely different from ours ( J. Proteome Res. 2018, 17 (10), 3606-3612. DOI: 10.1021/acs.jproteome.8b00620). Their study focused on the quantification of individual drug-metabolizing enzymes and transporters, whereas ours developed a targeted proteomics method to determine the allele-specific protein expression (ASPE) of a gene and advocated the use of the ASPE imbalance as the phenotype for identifying cis-regulatory genetic variants of the gene. More importantly, the digestion enzyme trypsin interacts with three to four amino acid residues around scissile bonds, and certain residues, such as negatively charged amino acids, can significantly affect the digestion efficiency. The QconCAT standard reported in our study differs from conventional QconCAT standards such as that used by Russell et al. in that at least 15 native flanking amino acids were included to ensure accurate measurement of ASPE ratios.