title: Purification and analysis of lung and plasma angiotensin I-converting enzyme by high-performance liquid chromatography.
abstract: We purified angiotensin I-converting enzyme (ACE) from pig and human lung and plasma for comparison of some physicochemical properties between the endothelial membrane-bound form and the soluble form of the enzyme. After affinity chromatography on Sepharose CL-4B/lisinopril, gel-filtration HPLC on Superose 12 achieved homogeneity for both forms as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Whatever the source of ACE, the molecular weight was 300 +/- 40 kDa after calibration of Superose 12 with standard globular proteins and 172 +/- 4 kDa by SDS-PAGE, with or without reduction, a result suggesting interactions between the glycopolypeptide chain and the chromatographic gel possibly related to the overall shape and sugar content of the enzyme. Ion-exchange HPLC analysis on TSK-DEAE showed that the membrane-bound and soluble forms of ACE are not isoenzymes, although isoelectrofocusing did show that the isoelectric point of soluble ACE was lower than those of tissue ACE, suggesting a different glycosylation. No significant difference between porcine and human ACE appeared. HPLC methods seem to be of particular interest for the purification of ACE with a high yield and for the analysis of its putative differently glycosylated isoforms.
authors.affiliation: Laboratoire de Chimie Biologique, UFR Pharmacie-Paris V, France.
authors.forename: B
authors.initials: B
authors.lastname: Baudin
authors.forename: A
authors.initials: A
authors.lastname: Tahraoui
authors.forename: F C
authors.initials: FC
authors.lastname: Baumann
authors.forename: D
authors.initials: D
authors.lastname: Robic
authors.forename: L
authors.initials: L
authors.lastname: Drouet
authors.forename: Y
authors.initials: Y
authors.lastname: Legrand
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002852:Chromatography, Ion Exchange; D006031:Glycosylation; D006801:Humans; D007526:Isoelectric Point; D008168:Lung / Q000201:enzymology*; D008970:Molecular Weight; D007703:Peptidyl-Dipeptidase A / Q000097:blood / Q000737:chemistry / Q000302:isolation & purification*; D013045:Species Specificity; D013552:Swine
chemical_list: D007703:Peptidyl-Dipeptidase A