title: The distal ectodomain of angiotensin-converting enzyme regulates its cleavage-secretion from the cell surface.
abstract: Angiotensin-converting enzyme (ACE) is a type I ectoprotein that is cleaved off the cell surface by a plasma membrane-bound metalloprotease. However, CD4, another type I ectoprotein does not undergo such cleavage-secretion. In this study, we investigated the structural determinants of the ACE protein that regulate the cleavage-secretion process. Substitution and deletion mutations revealed that the cytoplasmic domain, the transmembrane domain, and the juxtamembrane region encompassing the major and the minor cleavage sites of ACE do not regulate its cleavage. Moreover, a chimeric protein containing the distal extracellular domain of CD4 and the juxtamembrane, transmembrane, and the cytoplasmic domains of ACE, although transported to the cell surface, was not cleavage-secreted. In contrast, the distal extracellular domain of ACE was shown to be the important determinant: a protein containing the distal extracellular domain of ACE and the juxtamembrane, transmembrane, and cytoplasmic domain of CD4 was efficiently cleaved off the cell surface. The chimeric protein was cleaved within the CD4 sequence and the responsible enzymatic activity was inhibited by Compound 3, a relatively specific inhibitor of the ACE secretase activity. These results demonstrate that, in a chimeric protein, the distal extracellular domain of a cleavable protein, such as ACE, can induce a proteolytic cleavage within the juxtamembrane domain of an uncleaved protein such as CD4.
authors.affiliation: Department of Molecular Cardiology, Lerner Research Institute, The Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
authors.forename: R
authors.initials: R
authors.lastname: Sadhukhan
authors.forename: G C
authors.initials: GC
authors.lastname: Sen
authors.forename: R
authors.initials: R
authors.lastname: Ramchandran
authors.forename: I
authors.initials: I
authors.lastname: Sen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015704:CD4 Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003593:Cytoplasm / Q000201:enzymology; D005109:Extracellular Matrix / Q000201:enzymology; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D007703:Peptidyl-Dipeptidase A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013499:Surface Properties; D013737:Testis / Q000201:enzymology*
chemical_list: D015704:CD4 Antigens; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D007703:Peptidyl-Dipeptidase A