Please use this identifier to cite or link to this item:
Type: Artigo de periódico
Title: New mass spectrometric assay for angiotensin-converting enzyme 2 activity
Author: Elased, KM
Cunha, TS
Gurley, SB
Coffman, TM
Morris, M
Abstract: A novel assay was developed for evaluation of mouse angiotensin- converting enzyme ( ACE) 2 and recombinant human ACE2 ( rACE2) activity. Using surface- enhanced laser desorption/ ionization time of flight mass spectrometry ( MS) with ProteinChip Array technology, ACE1 and ACE2 activity could be measured using natural peptide substrates. Plasma from C57BL/ 6 mice, kidney from wild- type and ACE2 knockout mice, and rACE2 were used for assay validation. Plasma or tissue extracts were incubated with angiotensin I ( Ang I; 1296 m/ z) or angiotensin II ( Ang II; 1045 m/ z). Reaction mixtures were spotted onto the ProteinChips WCX2 and peptides detected using surface- enhanced laser desorption/ ionization time of flight MS. MS peaks for the substrates, Ang I and Ang II, and the generated peptides, Ang ( 1- 7) and Ang ( 1- 9), were monitored. The ACE2 inhibitor MLN 4760 ( 0.01 to 100 mu mol/ L) significantly inhibited rACE2 activity ( IC50 = 3 nmol/ L). Ang II was preferably cleaved by rACE2 ( km = 5 mu mol/ L), whereas Ang I was not a good substrate for rACE2. There was no detectable ACE2 activity in plasma. Assay specificity was validated in a model of ACE2 gene deletion. In kidney extract from ACE2- deficient mice, there was no generation of Ang ( 1- 7) from Ang II. However, Ang ( 1- 7) was produced when Ang I was used as a substrate. In conclusion, we developed a specific and sensitive assay for ACE2 activity, which used the natural endogenous peptide substrate Ang II. This approach allows for the rapid screening for ACE2, which has applications in drug testing, high- throughput enzymatic assays, and identification of novel substrates/ inhibitors of the renin- angiotensin system.
Subject: angiotensin
renin-angiotensin system
angiotensin converting enzyme
Country: EUA
Editor: Lippincott Williams & Wilkins
Citation: Hypertension. Lippincott Williams & Wilkins, v. 47, n. 5, n. 1010, n. 1017, 2006.
Rights: fechado
Identifier DOI: 10.1161/01.HYP.0000215588.38539.30
Date Issue: 2006
Appears in Collections:Unicamp - Artigos e Outros Documentos

Files in This Item:
File Description SizeFormat 
WOS000236943400034.pdf568.93 kBAdobe PDFView/Open

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.