Angiotensin-converting enzyme of the human small intestine. Subunit and quaternary structure, biosynthesis and membrane association.

摘要: Angiotensin-converting enzyme (ACE) was isolated from detergent-derived extracts of human intestinal brush-border membranes (BBMs) by immunoprecipitation using a monoclonal antibody. Analysis the immunoprecipitates SDS/PAGE revealed polypeptide apparent M(r) 184,000 under reducing and non-reducing conditions, indicating that ACE does not contain intermolecular disulphide bridges. The quaternary structure examined cross-linking experiments with dithiobis[succinimidylpropionate] (DSP) density gradient centrifugation on sucrose gradients. Both approaches demonstrated is assembled in membrane as monomer. By contrast, control glycoprotein aminopeptidase N (ApN) exists dimer. Biosynthetic labelling tissue explants 184,000-M(r) protein generated single-polypeptide, mannose-rich precursor (M(r) 175,000) modification carbohydrate side-chains Golgi apparatus. mode association mature form BBMs investigated hydrophobic right-side-out vesicles photoactivatable carbene-generating reagent 125I-labelled 3-(trifluoromethyl)-3-(m[formylamino]phenyl)diazirine (125I-labelled TID), followed treatment trypsin at dilutions do cause substantial degradation ACE. These studies associated via segment. Furthermore, 35S-labelled inside-out possesses cytoplasmic tail, therefore has transmembraneous orientation.