RT Journal Article SR Electronic T1 QUANTITATIVE ANALYSIS OF CYTOCHROME P450 ISOZYMES BY MEANS OF UNIQUE ISOZYME-SPECIFIC TRYPTIC PEPTIDES: A PROTEOMIC APPROACH JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP 1399 OP 1407 DO 10.1124/dmd.105.004812 VO 33 IS 9 A1 Michail A. Alterman A1 Boris Kornilayev A1 Tatyana Duzhak A1 Dmitry Yakovlev YR 2005 UL http://dmd.aspetjournals.org/content/33/9/1399.abstract AB A novel matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry method has been developed to quantitate cytochrome P450 (P450) isozymes based on their unique isozyme-specific tryptic peptides. It was shown that the molar ratio of P450 isozyme-specific peptides is linearly proportional to the mass peak area ratio of corresponding peptides not only in simple two-peptide mixtures, but also in complex digest mixtures. This approach is applicable both to in-gel (as shown for CYP2B1 and CYP2B2) and in-solution digests (as shown for CYP1A2, CYP2E1, and CYP2C19) and does not require introduction of stable isotopes or labeling with isotope-coded affinity tagging. The relative and absolute quantitation can be performed after developing corresponding calibration curves with synthesized P450 isozyme-specific peptide standards. The absolute quantitation of human P450 isozymes was performed by using CYP2B2 isozyme-specific peptide (1306.7 Da) as the universal internal standard. The utility of this approach was demonstrated for two highly homologous (>97%) rat liver CYP2B1 and CYP2B2 and three human P450 isozymes belonging to two different families and three different subfamilies: CYP1A2, CYP2E1, and CYP2C19. In summary, we have demonstrated that MALDI TOF-based peptide mass fingerprinting of different cytochrome P450 isozymes can provide not only qualitative but quantitative data, too. The American Society for Pharmacology and Experimental Therapeutics