SEPARATION AND PURIFICATION OF N-DOMAIN OF BOVINE LUNG ANGIOTENSIN I-CONVERTING ENZYME BY SIZE EXCLUSION CHROMATOGRAPHY

Abstract

Bovine lung angiotensin I-converting enzyme is a monomer with two active sites, in its two (N and C) homologous domains. A process is described for the preparative isolation of the ACE N-domain to either the ACE soluble monomer or ACE aggregated one. After exposure to moderate heat for 7 hours and in presence of a protease, ACE N-domain was obtained as a whole, and only fragments of the C-domain. N-domain purified was separated by Sephacryl S-300 HR chromatography, and a recovery of 80% was obtained. Molecular mass was estimated to be 100 kDa by sodium dodecyl sulfate gel electrophoresis. After purifying and partially sequencing this domain, we have investigated some catalytic properties and the inhibition by captopril.

ACKNOWLEDGMENTS

This research was supported by grants PM97-0003 and PB96-1446 from DGICYT, Ministerio de Educación y Cultura, Spain.