Linked Life Data

18792928

Source:http://linkedlifedata.com/resource/pubmed/id/18792928

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
20
pubmed:dateCreated
2008-12-16
pubmed:abstractText
The human cytochrome P450 (P450) superfamily consists of membrane-bound proteins that metabolize a myriad of xenobiotics and endogenous compounds. Quantification of P450 expression in various tissues under normal and induced conditions has an important role in drug safety and efficacy. Conventional immunoquantification methods have poor dynamic range, low throughput, and a limited number of specific antibodies. Recent advances in MS-based quantitative proteomics enable absolute protein quantification in a complex biological mixture. We have developed a gel-free MS-based protein quantification strategy to quantify CYP3A enzymes in human liver microsomes (HLM). Recombinant protein-derived proteotypic peptides and synthetic stable isotope-labeled proteotypic peptides were used as calibration standards and internal standards, respectively. The lower limit of quantification was approximately 20 fmol P450. In two separate panels of HLM examined (n = 11 and n = 22), CYP3A, CYP3A4 and CYP3A5 concentrations were determined reproducibly (CV <or=27%). The MS-based method strongly correlated with the immunoquantification method (r(2)>or=0.87) and marker activities (r(2)>or=0.88), including testosterone 6beta-hydroxylation (CYP3A), midazolam 1'-hydroxylation (CYP3A), itraconazole 6-hydroxylation (CYP3A4) and CYP3A5-mediated vincristine M1 formation (CYP3A5). Taken together, our MS-based method provides a specific, sensitive and reliable means of P450 protein quantification and should facilitate P450 characterization during drug development, especially when specific substrates and/or antibodies are unavailable.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
1615-9861
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
8
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
4186-96
pubmed:dateRevised
2010-11-18
pubmed:meshHeading
pubmed:year
2008
pubmed:articleTitle
A gel-free MS-based quantitative proteomic approach accurately measures cytochrome P450 protein concentrations in human liver microsomes.
pubmed:affiliation
School of Pharmacy, The University of North Carolina at Chapel Hill, NC 27599, USA. michael.wang@unc.edu
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't