scholarly journals Metabolism of neuropeptides. Hydrolysis of the angiotensins, bradykinin, substance P and oxytocin by pig kidney microvillar membranes

1987 ◽  
Vol 241 (1) ◽  
pp. 237-247 ◽  
Author(s):  
S L Stephenson ◽  
A J Kenny
Keyword(s):  
Substance P ◽  
Membrane Fraction ◽  
Angiotensin I ◽  
Peptide Bonds ◽  
Prolonged Incubation ◽  
Pig Kidney ◽  
Angiotensin I Converting Enzyme ◽  
Endopeptidase 24.11 ◽  
20 Nm ◽  
Hydrolysis Of

Microvillar membranes derived from the brush border of the renal proximal tubule are very rich in peptidases. Pig kidney microvilli contain endopeptidase-24.11 associated with a battery of exopeptidases. The manner by which some neuropeptides are degraded by the combined attack of the peptidases of this membrane has been investigated. The contribution of individual peptidases was assessed by including inhibitors (phosphoramidon, captopril, amastatin and di-isopropyl fluorophosphate) with the membrane fraction when incubated with the peptides. Substance P, bradykinin and angiotensins I, II and III and insulin B-chain were rapidly hydrolysed by kidney microvilli. Oxytocin was hydrolysed much more slowly, but no products were detected from [Arg8]vasopressin or insulin under the conditions used for other peptides. The peptide bonds hydrolysed were identified and the contributions of the different peptidases were quantified. For each of the susceptible peptides, the main contribution came from endopeptidase-24.11 (inhibited by phosphoramidon). Peptidyl dipeptidase A (angiotensin-I-converting enzyme) was of less importance, even in respect of angiotensin I and bradykinin. When [2,3-Pro3,4-3H]bradykinin was also investigated at a lower concentration (20 nM), the conclusions in regard to the contributions of the two peptidases were unchanged. The possibility that endopeptidase-24.11 might attack within the six-residue disulphide-bridged rings of oxytocin and vasopressin was examined by dansyl(5-dimethylaminonaphthalene-1-sulphonyl)ation and by reduction and carboxymethylation of the products after incubation. Additional peptides were only observed after prolonged incubation, consistent with hydrolysis at the Tyr2-Ile3 and Tyr2-Phe3 bonds respectively. These results show that a range of neuropeptides are efficiently degraded by microvillar membranes and that endopeptidase-24.11 plays a key role in this process.

scholarly journals The hydrolysis of α-human atrial natriuretic peptide by pig kidney microvillar membranes is initiated by endopeptidase-24.11

1987 ◽  
Vol 243 (1) ◽  
pp. 183-187 ◽  
Author(s):  
S L Stephenson ◽  
A J Kenny
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Angiotensin I ◽  
Human Atrial Natriuretic Peptide ◽  
Pig Kidney ◽  
Angiotensin I Converting Enzyme ◽  
Endopeptidase 24.11 ◽  
Hydrolysis Of ◽  
Atrial Natriuretic

alpha-Human atrial natriuretic peptide, a 28-amino-acid-residue peptide, was rapidly hydrolysed by pig kidney microvillar membranes in vitro, with a t1/2 of 8 min, comparable with the rate observed with angiotensins II and III. The products of hydrolysis were analysed by h.p.l.c., the pattern obtained with membranes being similar to that with purified endopeptidase-24.11 (EC 3.4.24.11). No hydrolysis by peptidyl dipeptidase A (angiotensin I converting enzyme, EC 3.4.15.1) was observed. The contribution of the various microvillar membrane peptidases was assessed by including specific inhibitors. Phosphoramidon, an inhibitor of endopeptidase-24.11, caused 80-100% suppression of the products. Captopril and amastatin (inhibitors of peptidyl dipeptidase A and aminopeptidases respectively) had no significant effect. Hydrolysis at an undefined site within the disulphide-linked ring occurred rapidly, followed by hydrolysis at other sites, including the Ser25--Phe26 bond.

scholarly journals The metabolism of neuropeptides. Neurokinin A (substance K) is a substrate for endopeptidase-24.11 but not for peptidyl dipeptidase A (angiotensin-converting enzyme)

1985 ◽  
Vol 231 (2) ◽  
pp. 357-361 ◽  
Author(s):  
N M Hooper ◽  
A J Kenny ◽  
A J Turner
Keyword(s):  
Substance P ◽  
Selective Inhibitor ◽  
Synaptic Membranes ◽  
Neurokinin A ◽  
Pig Kidney ◽  
Endopeptidase 24.11 ◽  
The Family ◽  
General Capacity ◽  
Substance K ◽  
Hydrolysis Of

Both endopeptidase-24.11 and peptidyl dipeptidase A have previously been shown to hydrolyse the neuropeptide substance P. The structurally related peptide neurokinin A is also shown to be hydrolysed by pig kidney endopeptidase-24.11. The identified products indicated hydrolysis at two sites, Ser5-Phe6 and Gly8-Leu9, consistent with the known specificity of the enzyme. The pattern of hydrolysis of neurokinin A by synaptic membranes prepared from pig striatum was similar to that observed with purified endopeptidase-24.11, and hydrolysis was substantially abolished by the selective inhibitor phosphoramidon. Peptidyl dipeptidase A purified from pig kidney was shown to hydrolyse substance P but not neurokinin A. It is concluded that endopeptidase-24.11 has the general capacity to hydrolyse and inactivate the family of tachykinin peptides, including substance P and neurokinin A.

scholarly journals A novel peptide-processing activity of insect peptidyl-dipeptidase A (angiotensin I-converting enzyme): the hydrolysis of lysyl-arginine and arginyl-arginine from the C-terminus of an insect prohormone peptide

1998 ◽  
Vol 330 (1) ◽  
pp. 61-65 ◽  
Author(s):  
R. Elwyn ISAAC ◽  
Liliane SCHOOFS ◽  
A. Tracy WILLIAMS ◽  
Dirk VEELAERT ◽  
Mohammed SAJID ◽  
...  
Keyword(s):  
Neurosecretory Cells ◽  
Single Domain ◽  
Converting Enzyme ◽  
Angiotensin I ◽  
Recognition Sequence ◽  
Angiotensin I Converting Enzyme ◽  
C Terminus ◽  
Processing Activity ◽  
Hydrolysis Of

Insect peptidyl-dipeptidase A [angiotensin I-converting enzyme (ACE)] is a soluble single-domain peptidyl-dipeptidase that has many properties in common with the C-domain of mammalian somatic ACE and with the single-domain mammalian germinal ACE. Mammalian somatic ACE is important in blood homoeostasis, but the role of ACE in insects is not known. Immunocytochemistry has been used to localize ACE in the neuroendocrine system of the locust, Locusta migratoria. Staining was observed in five groups of neurosecretory cells in the brain and suboesophageal ganglion, in the nervi corpori cardiaci, the storage part of the corpora cardiaca and in the nervi corpori allati. In three groups of neurosecretory cells, ACE co-localized with locustamyotropins, suggesting a possible role for the enzyme in the metabolism of these neuropeptides. We demonstrate in vitro a novel activity of ACE that removes pairs of basic amino acid residues from a locustamyotropin peptide extended at the C-terminus with either Gly-Lys-Arg or Gly-Arg-Arg, corresponding to a consensus recognition sequence for endoproteolysis of prohormone proteins by prohormone convertases. The low Km and high kcat values (Km 7.3 and 5.0 μM, kcat 226 and 207 s-1 for the hydrolysis of Phe-Ser-Pro-Arg-Leu-Gly-Lys-Arg and Phe-Ser-Pro-Arg-Leu-Gly-Arg-Arg, respectively) obtained for the hydrolysis of these two peptides by insect ACE means that these peptides, along with mammalian bradykinin, are the most favoured in vitro ACE substrates so far identified. The discovery of this in vitro prohormone-processing activity of insect ACE provides a possible explanation for the intracellular co-localization of the enzyme with locustamyotropin peptides, and provides evidence for a new role for ACE in the biosynthesis of peptide hormones and transmitters.

scholarly journals The metabolism of neuropeptides. Hydrolysis of peptides by the phosphoramidon-insensitive rat kidney enzyme ‘endopeptidase-2’ and by rat microvillar membranes

1988 ◽  
Vol 255 (1) ◽  
pp. 45-51 ◽  
Author(s):  
S L Stephenson ◽  
A J Kenny
Keyword(s):  
Substance P ◽  
Rat Kidney ◽  
Peptide Fragments ◽  
Hydrophobic Amino Acid ◽  
Prolonged Incubation ◽  
Minor Contribution ◽  
Kidney Enzyme ◽  
Hydrophobic Amino Acid Residue ◽  
A Minor ◽  
Hydrolysis Of

Endopeptidase-2, the second endopeptidase in rat kidney brush border [Kenny & Ingram (1987) Biochem. J. 245, 515-524] has been further characterized in regard to its specificity and its contribution to the hydrolysis of peptides by microvillar membrane preparations. The peptide products were identified, after incubating luliberin, substance P, bradykinin and angiotensins I, II and III with the purified enzyme. The bonds hydrolysed were those involving a hydrophobic amino acid residue, but this residue could be located at either the P1 or P1′ site. Luliberin was hydrolysed faster than other peptides tested, followed by substance P and bradykinin. Human alpha-atrial natriuretic peptide and the angiotensins were only slowly attacked. Oxytocin and [Arg8]vasopressin were not hydrolysed. No peptide fragments were detected on prolonged incubation with insulin, cytochrome c, ovalbumin and serum albumin. In comparison with pig endopeptidase-24.11 the rates for the susceptible peptides were, with the exception of luliberin, much lower for endopeptidase-2. Indeed, for bradykinin and substance P the ratio kcat./Km was two orders of magnitude lower. Since both endopeptidases are present in rat kidney microvilli, an assessment was made of the relative contributions to the hydrolysis of luliberin, bradykinin and substance P. Only for the first named was endopeptidase-2 the dominant enzyme; for bradykinin it made an equal, and for substance P a minor, contribution.

Cleavage of des-Arg9-bradykinin by angiotensin I-converting enzyme from pig kidney cortex

1985 ◽  
Vol 41 (3) ◽  
pp. 325-328 ◽  
Author(s):  
G. Oshima ◽  
Y. Hiraga ◽  
K. Shirono ◽  
S. Oh-ishi ◽  
S. Sakakibara ◽  
...  
Keyword(s):  
Kidney Cortex ◽  
Converting Enzyme ◽  
Angiotensin I ◽  
Pig Kidney ◽  
Angiotensin I Converting Enzyme

scholarly journals Hydrolysis of human and pig brain natriuretic peptides, urodilatin, C-type natriuretic peptide and some C-receptor ligands by endopeptidase-24.11

1993 ◽  
Vol 291 (1) ◽  
pp. 83-88 ◽  
Author(s):  
A J Kenny ◽  
A Bourne ◽  
J Ingram
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Brain Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Natriuretic Peptides ◽  
Receptor Ligands ◽  
Prolonged Incubation ◽  
Endopeptidase 24.11 ◽  
Pig Brain ◽  
Hydrolysis Of ◽  
Atrial Natriuretic

Endopeptidase-24.11 (E-24.11, EC 3.4.24.11) is widely believed to play a physiological role in metabolizing atrial natriuretic peptide (ANP). Since the discovery of ANP, new natriuretic peptides have been isolated and other peptides synthesized as receptor ligands. The hydrolysis in vitro of six related peptides by the endopeptidase has been studied, mainly by h.p.l.c. The initial attack on the 32-residue form of pig brain natriuretic peptide (pBNP-32) was shown to be at the Ser20-Leu21 bond, as had been previously shown for the 26-residue form. In contrast, human brain natriuretic peptide-32 (hBNP-32), which differs in ten residues from pBNP-32, was attacked first at the Met4-Val5 bond, releasing the N-terminal tetrapeptide, and only later at bonds within the ring: at Arg17-Ile18 and subsequently at four other sites. Urodilatin, which has a four-residue extension at the N-terminus compared with alpha-human atrial natriuretic peptide-28 (alpha-hANP), was degraded at about half the rate of the latter, though the C-terminal Phe-Arg-Tyr was released at the same rate. The 22-residue C-type natriuretic peptide was hydrolysed more rapidly than alpha-hANP, as were two C-receptor ligands (peptides with deletions within the ring): C-ANP4-23 (rANP4-23 des-Gln18,Ser19,Gly20,Leu21,Gly22) and SC 46542 (hANP5-28 des-Phe8,Gly9,Ala17,Gln18). Angiotensin-converting enzyme failed to hydrolyse pBNP-32, hBNP-32 or 125I-rat (r) ANP, even after prolonged incubation. Km and kcat values were determined for the hydrolysis of alpha-hANP, porcine BNP-26, porcine BNP-32 and 125I-rANP by E-24.11. Ki values were determined for six peptides, alpha-hANP, urodilatin, hBNP-32, C-type natriuretic peptide (CNP), SC 46542 and C-type natriuretic peptide (C-ANP4-23), in radiometric assays of E-24.11 with either [125I] insulin B chain or [125I] rANP as substrate. The Ki values (2.5-13 microM) for CNP were the lowest of any of the group, whereas those for hBNP-32 (151-172 microM) were the highest. The physiological significance of these results is discussed, especially in regard to the relative resistance of hBNP-32 to attack and the ability of the C-receptor ligands to compete with natriuretic peptides for hydrolysis by E-24.11.

Sign in / Sign up

Export Citation Format

Share Document