Pharmaceutical Sciences Faculty Publications
Optimized Method for Computing (18)O/(16)O Ratios of Differentially Stable-Isotope Labeled Peptides in the Context of Postdigestion (18)O Exchange/Labeling
Document Type
Article
Publication Date
7-1-2010
Journal Title
Analytical Chemistry
ISSN
1520-6882
Volume
82
Issue
13
First Page
5878
Last Page
5886
DOI
10.1021/ac101284c
PubMed ID
20540505
PubMed Central® ID
PMC3479679
Abstract
Differential (18)O/(16)O stable isotope labeling of peptides that relies on enzyme-catalyzed oxygen exchange at their carboxyl termini in the presence of H(2)(18)O has been widely used for relative quantitation of peptides/proteins. The role of tryptic proteolysis in bottom-up shotgun proteomics and low reagent costs have made trypsin-catalyzed (18)O postdigestion exchange a convenient and affordable stable isotope labeling approach. However, it is known that trypsin-catalyzed (18)O exchange at the carboxyl terminus is in many instances inhomogeneous/incomplete. The extent of the (18)O exchange/incorporation fluctuates from peptide to peptide mostly due to variable enzyme-substrate affinity. Thus, accurate calculation and interpretation of peptide ratios are analytically complicated and in some regard deficient. Therefore, a computational approach capable of improved measurement of actual (18)O incorporation for each differentially labeled peptide pair is needed. In this regard, we have developed an algorithmic method that relies on the trapezoidal rule to integrate peak intensities of all detected isotopic species across a particular peptide ion over the retention time, which fits the isotopic manifold to Poisson distributions. Optimal values for manifold fitting were calculated and then (18)O/(16)O ratios derived via evolutionary programming. The algorithm is tested using trypsin-catalyzed (18)O postdigestion exchange to differentially label bovine serum albumin (BSA) at a priori determined ratios. Both accuracy and precision are improved utilizing this rigorous mathematical approach. We further demonstrate the effectiveness of this method to accurately calculate (18)O/(16)O ratios in a large scale proteomic quantitation of detergent resistant membrane microdomains (DRMMs) isolated from cells expressing wild-type HIV-1 Gag and its nonmyristylated mutant.
Keywords
Algorithms, amino acid sequence, isotope labeling, oxygen isotopes, peptides, human immunodeficiency virus
Recommended Citation
Ye, Xiaoying; Luke, Brian T.; Johann, Donald J.; Ono, Akira; Prieto, Darue A.; Chan, King C.; Issaq, Haleem J.; Veenstra, Timothy D.; and Blonder, Josip, "Optimized Method for Computing (18)O/(16)O Ratios of Differentially Stable-Isotope Labeled Peptides in the Context of Postdigestion (18)O Exchange/Labeling" (2010). Pharmaceutical Sciences Faculty Publications. 269.
https://digitalcommons.cedarville.edu/pharmaceutical_sciences_publications/269