Insulin stimulates cell surface aminopeptidase activity toward vasopressin in adipocytes

Abstract

We previously discovered that insulin stimulates the marked translocation of a novel membrane aminopeptidase, designated vp165 for vesicle protein of 165 kDa, to the cell surface in adipocytes. To examine the hypothesis that this enzyme acts on peptide hormones, we assessed the relative affinity of the enzyme for 22 peptide hormones by measuring the inhibitory effect of each on the hydrolysis of a fluorogenic substrate, and we directly assayed the cleavage of four of these. Angiotensin III, angiotensin IV, and Lys-bradykinin bound to the enzyme with half-saturation constants between 20 and 600 nM and were cleaved by vp165. Vasopressin bound with lower affinity but at saturation was cleaved more rapidly. Subsequently, the effect of insulin on the rates of cleavage of 125I-labeled vasopressin by intact 3T3-L1 and rat adipocytes was determined. With both cell types, vasopressin cleavage was stimulated approximately threefold. These findings indicate that a physiological role for vp165 may be the processing of peptide hormones and that insulin could enhance the cleavage of extracellular substrates by eliciting the translocation of vp165 to the cell surface.

International Union of Basic and Clinical Pharmacology. XCIX. Angiotensin Receptors: Interpreters of Pathophysiological Angiotensinergic Stimuli

“…AngIV and divalinal display competitive kinetics, indicating that AngIV binding site ligands mediate their peptidase inhibitor effects by binding to the catalytic site of IRAP . IRAP is known to play a role in the in vivo metabolism of vasopressin and Lys-bradykinin bound to the enzyme (Herbst et al, 1997;Wallis et al, 2007). Substrates identified in vitro for IRAP include vasopressin (AVP), oxytocin, somatostatin, Leu-and Metenkephalin, and Lys-bradykinin (Herbst et al, 1997;Matsumoto et al, 2001;Lew et al, 2003).…”

“…In vivo, IRAP hydrolyzes the N-terminal amino acid from neuropeptides including arginine-vasopressin, cholecystokinin-8, dynorphin, met-enkephalin, lysine-bradykinin, neurokinin A, neuromedin B, oxytocin and somatastatin, (Herbst et al, 1997;Matsumoto et al, 2001;Lew et al, 2003). Accordingly, IRAP was proposed as the first AngIV binding site (see Fig.…”

Structure-Activity Study of LVV-Hemorphin-7: Angiotensin AT4 Receptor Ligand and Inhibitor of Insulin-Regulated Aminopeptidase

“…The enzyme has previously been defined as specifically cleaving the N-terminal amino acid CysXaa-, in which the halfcystine residue is involved in a disulfide loop, notably in oxytocin, vasopressin, and somatostatin (Herbst et al, 1997). N-Terminal cysteine residues seem to be the preferential targets for the enzyme; however, other peptides that possess N-terminal cysteine residues and intramolecular disulfide bonds, such as calcitonin and endothelin, are not cleaved by the enzyme.…”

“…N-Terminal cysteine residues seem to be the preferential targets for the enzyme; however, other peptides that possess N-terminal cysteine residues and intramolecular disulfide bonds, such as calcitonin and endothelin, are not cleaved by the enzyme. Other peptides that are readily cleaved by IRAP include Lys-bradykinin, met-enkephalin, dynorphin A, neurokinin A, and neuromedin B (Herbst et al, 1997), which possess a range of N-terminal residues. In contrast to AT 4 ligands the affinities of such substrates are in the mid-micromolar range, as is common with peptidases.…”

AT4 receptor binding in the developing rabbit

“…The authors contend that the myriad physiological effects of AT 4 ligands are due, in fact, to their unique ability to inhibit the peptidase activity of IRAP competitively, thus potentiating the actions of endogenous peptides that are normally degraded by IRAP. This notion is supported by the observation that AngIV, acting as a substrate for IRAP, has an unusually low K m of 20 nM (Herbst et al, 1997). While at first glance, this may seem a reasonable explanation for the actions of AT 4 receptor ligands, many published and unpublished data regarding AT 4 receptor function seem inconsistent with this explanation.…”