Regulated cleavage-secretion of the membrane-bound angiotensin-converting enzyme.

Angiotensin-converting enzyme (ACE; EC 3.4.1.15.1) exists in both membrane-bound and soluble forms. Phase separation in Triton X-114 and a competitive e.l.i.s.a. have been employed to characterize the activity which post-translationally converts the amphipathic, membrane-bound form of ACE in pig kidney microvilli into a hydrophilic, soluble form. This secretase activity was enriched to a similar extent as other microvillar membrane proteins, was tightly membrane-associated, being resistant to extensive washing of the microvillar membranes with 0.5 M NaCl, and displayed a pH optimum of 8.4. The ACE secretase was not affected by inhibitors of serine-, thiol- or aspartic-proteases, nor by reducing agents or alpha 2-macroglobulin. The metal chelators, EDTA and 1,10-phenanthroline, inhibited the secretase activity, with, in the case of EDTA, an inhibitor concentration of 2.5 mM causing 50% inhibition. In contrast, EGTA inhibited the secretase by a maximum of 15% at a concentration of 10 mM. The inhibition of EDTA was reactivated substantially (83%) by Mg2+ ions, and partially (34% and 29%) by Zn2+ and Mn2+ ions respectively. This EDTA-sensitive secretase activity was also present in microsomal membranes prepared from pig lung and testis, and from human lung and placenta, but was absent from human kidney and human and pig intestinal brush-border membranes. The form of ACE released from the microvillar membrane by the secretase co-migrated on SDS/PAGE with ACE purified from pig plasma, thus the action and location of the secretase would be consistent with it possibly having a role in the post-translational proteolytic cleavage of membrane-bound ACE to generate the soluble form found in blood, amniotic fluid, seminal plasma and other body fluids.

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Characterization of a membrane-bound angiotensin-converting enzyme isoform in crayfish testis and evidence for its release into the seminal fluid

FEBS Journal ◽

10.1111/j.1742-4658.2009.07169.x ◽

2009 ◽

Vol 276 (17) ◽

pp. 4727-4738 ◽

Cited By ~ 3

Author(s):

Juraj Simunic ◽

Daniel Soyez ◽

Nédia Kamech

Keyword(s):

Angiotensin Converting Enzyme ◽

Seminal Fluid ◽

Converting Enzyme ◽

Membrane Bound

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Angiotensin converting enzyme 2 is a novel target of the γ-secretase complex

10.1101/2020.09.01.277954 ◽

2020 ◽

Author(s):

Alberto Bartolomé ◽

Jiani Liang ◽

Pengfei Wang ◽

David D. Ho ◽

Utpal B. Pajvani

Keyword(s):

Angiotensin Converting Enzyme ◽

Renin Angiotensin System ◽

Structural Similarity ◽

Ectodomain Shedding ◽

Converting Enzyme ◽

Angiotensin System ◽

Angiotensin Converting Enzyme 2 ◽

Membrane Bound ◽

Novel Target ◽

Functional Receptor

AbstractAngiotensin converting enzyme 2 (ACE2) is a key regulator of the renin-angiotensin system, but also the functional receptor of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Based on structural similarity with other γ-secretase (γS) targets, we hypothesized that ACE2 may be affected by γS proteolytic activity. We found that after ectodomain shedding, ACE2 is targeted for intramembrane proteolysis by γS, releasing a soluble ACE2 C-terminal fragment. Consistently, chemical or genetic inhibition of γS results in the accumulation of a membrane-bound fragment of ectodomain-deficient ACE2. Although chemical inhibition of γS does not alter SARS-CoV-2 cell entry, these data point to a novel pathway for cellular ACE2 trafficking.

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Interaction of SARS-CoV-2 and Other Coronavirus With ACE (Angiotensin-Converting Enzyme)-2 as Their Main Receptor

Hypertension ◽

10.1161/hypertensionaha.120.15256 ◽

2020 ◽

Vol 76 (5) ◽

pp. 1339-1349 ◽

Cited By ~ 4

Author(s):

Anne M. Davidson ◽

Jan Wysocki ◽

Daniel Batlle

Keyword(s):

Angiotensin Converting Enzyme ◽

Scientific Research ◽

Converting Enzyme ◽

Angiotensin Converting Enzyme 2 ◽

Therapeutic Implications ◽

Protein Receptor ◽

Type Ii Pneumocytes ◽

A Cell ◽

Membrane Bound ◽

Novel Coronavirus

Severe acute respiratory syndrome coronavirus (SARS-CoV)-2 originated from Wuhan, China, in December 2019 and rapidly spread to other areas worldwide. Since then, coronavirus disease 2019 (COVID-19) has reached pandemic proportions with >570 000 deaths globally by mid-July 2020. The magnitude of the outbreak and the potentially severe clinical course of COVID-19 has led to a burst of scientific research on this novel coronavirus and its host receptor ACE (angiotensin-converting enzyme)-2. ACE2 is a homolog of the ACE that acts on several substrates in the renin-Ang (angiotensin) system. With unprecedented speed, scientific research has solved the structure of SARS-CoV-2 and imaged its binding with the ACE2 receptor. In SARS-CoV-2 infection, the viral S (spike) protein receptor-binding domain binds to ACE2 to enter the host cell. ACE2 expression in the lungs is relatively low, but it is present in type II pneumocytes—a cell type also endowed with TMPRSS2 (transmembrane protease serine 2). This protease is critical for priming the SARS-CoV-2 S protein to complex with ACE2 and enter the cells. Herein, we review the current understanding of the interaction of SARS-CoV-2 with ACE2 as it has rapidly unfolded over the last months. While it should not be assumed that we have a complete picture of SARS-CoV-2 mechanism of infection and its interaction with ACE2, much has been learned with clear therapeutic implications. Potential therapies aimed at intercepting SARS-CoV-2 from reaching the full-length membrane-bound ACE2 receptor using soluble ACE2 protein and other potential approaches are briefly discussed as well.

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Angiotensin converting enzyme 2 is a novel target of the γ-secretase complex

Scientific Reports ◽

10.1038/s41598-021-89379-x ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Alberto Bartolomé ◽

Jiani Liang ◽

Pengfei Wang ◽

David D. Ho ◽

Utpal B. Pajvani

Keyword(s):

Angiotensin Converting Enzyme ◽

Renin Angiotensin System ◽

Structural Similarity ◽

Ectodomain Shedding ◽

Converting Enzyme ◽

Angiotensin System ◽

Angiotensin Converting Enzyme 2 ◽

Membrane Bound ◽

Novel Target ◽

Functional Receptor

AbstractAngiotensin converting enzyme 2 (ACE2) is a key regulator of the renin-angiotensin system, but also the functional receptor of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Based on structural similarity with other γ-secretase (γS) targets, we hypothesized that ACE2 may be affected by γS proteolytic activity. We found that after ectodomain shedding, ACE2 is targeted for intramembrane proteolysis by γS, releasing a soluble ACE2 C-terminal fragment. Consistently, chemical or genetic inhibition of γS results in the accumulation of a membrane-bound fragment of ectodomain-deficient ACE2. Although chemical inhibition of γS does not alter SARS-CoV-2 cell entry, these data point to a novel pathway for cellular ACE2 trafficking.

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Melatonin, cardiovascular disease and COVID-19: A potential therapeutic strategy?

Melatonin Research ◽

10.32794/mr11250065 ◽

2020 ◽

Vol 3 (3) ◽

pp. 318-321 ◽

Cited By ~ 3

Author(s):

Alberto Dominguez-Rodriguez ◽

Pedro Abreu-Gonzalez ◽

Paul E Marik ◽

Russel J Reiter

Keyword(s):

Reactive Oxygen Species ◽

Cardiovascular Disease ◽

Endothelial Dysfunction ◽

Angiotensin Converting Enzyme ◽

Therapeutic Strategy ◽

Oxygen Species ◽

Converting Enzyme ◽

Angiotensin Converting Enzyme 2 ◽

Ros Formation ◽

Membrane Bound

The mechanism for SARS-CoV-2 infection is the requisite binding of the virus to the membrane-bound form of angiotensin-converting enzyme 2 (ACE2) and internalization of the complex by the host cell. SARS-CoV-2 induced endothelial dysfunction and cardiovascular injury are probably initiated by increases in the phosphorylation levels of JAK2 and STAT3 and resultant reactive oxygen species (ROS) formation. These pathological alterations are speculated to be strikingly reversed by melatonin

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Human small intestinal angiotensin-converting enzyme: intracellular transport, secretion and glycosylation

Biochemical Journal ◽

10.1042/bj2960607 ◽

1993 ◽

Vol 296 (3) ◽

pp. 607-615 ◽

Cited By ~ 14

Author(s):

H Y Naim

Keyword(s):

Angiotensin Converting Enzyme ◽

Molecular Mass ◽

Brush Border ◽

Intracellular Transport ◽

Apparent Molecular Mass ◽

Intestinal Cells ◽

Converting Enzyme ◽

Human Biopsy ◽

Membrane Bound ◽

Small Intestinal

Human intestinal angiotensin-converting enzyme (ACE) exists in the brush-border membrane as a monomeric protein of apparent molecular mass 184 kDa. It is associated with the membrane via a hydrophobic segment and has a transmembrane orientation [Naim (1992) Biochem. J. 286, 451-457]. In addition to the membrane-bound form (ACEm), hydrophilic forms of ACE (ACEsec) can be identified in biosynthetically labelled intestinal cells. Thus the culture medium of biosynthetically labelled human biopsy samples contains an ACE molecule which has an apparent molecular mass similar to that of its membrane-bound counterpart. The secreted ACEsec forms follow a precursor/product relationship with the mature ACE molecule. The effect of the monomeric structure of ACE in its intracellular transport and secretion was investigated by pulse-chase experiments on human biopsy samples labelled with [35S]methionine. The results reveal 2-3-fold slower transport of ACE from the endoplasmic reticulum (ER) to the Golgi as compared with the homodimeric proteins dipeptidylpeptidase IV and aminopeptidase N. Further, the transport kinetics of ACE are comparable with those of human sucrase-isomaltase and human maltase-glucoamylase, two brush-border disaccharidases that do not form homodimers in the ER of human small-intestinal cells. These findings strongly suggest that homodimerization of brush-border proteins may influence the rate of transport of these proteins from the ER to the Golgi. The effect of glycosylation on the transport and secretion of ACE was investigated by utilizing several inhibitors of glycan processing. Here, secretion of ACE molecules continued to take place, albeit to a considerably lesser extent. In fact, approx. 2-fold less ACE molecules were secreted in the presence of inhibitors of ER glucosidases I and II and cis-Golgi mannosidase-I, suggesting that carbohydrate processing is important in the attainment of a transport-competent conformation.

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Human recombinant membrane-bound aminopeptidase P: production of a soluble form and characterization using novel, internally quenched fluorescent substrates

Biochemical Journal ◽

10.1042/bj20040849 ◽

2005 ◽

Vol 385 (2) ◽

pp. 389-397 ◽

Cited By ~ 25

Author(s):

Giuseppe MOLINARO ◽

Adriana K. CARMONA ◽

Maria A. JULIANO ◽

Luiz JULIANO ◽

Elena MALITSKAYA ◽

...

Keyword(s):

Amino Acid ◽

Angiotensin Converting Enzyme ◽

Angiotensin Converting Enzyme Inhibitors ◽

Polyclonal Antiserum ◽

Soluble Form ◽

Converting Enzyme ◽

Converting Enzyme Inhibitors ◽

Fluorescent Peptide ◽

Membrane Bound ◽

Aminopeptidase P

APP (aminopeptidase P) has the unique ability to cleave the N-terminal amino acid residue from peptides exhibiting a proline at P1′. Despite its putative involvement in the processing of bioactive peptides, among them the kinins, little is known about the physiological roles of both human forms of APP. The purpose of the present study is first to engineer and characterize a secreted form of hmAPP (human membrane-bound APP). Our biochemical analysis has shown that the expressed glycosylated protein is fully functional, and exhibits enzymic parameters similar to those described previously for mAPP purified from porcine or bovine lungs or expressed from a porcine clone. This soluble form of hmAPP cross-reacts with a polyclonal antiserum raised against a 469-amino-acid hmAPP fragment produced in Escherichia coli. Secondly, we synthesized three internally quenched fluorescent peptide substrates that exhibit a similar affinity for the enzyme than its natural substrates, the kinins, and a higher affinity compared with the tripeptide Arg-Pro-Pro used until now for the quantification of APP in biological samples. These new substrates represent a helpful analytical tool for rapid and reliable screening of patients susceptible to adverse reactions associated with angiotensin-converting enzyme inhibitors or novel vasopeptidase (mixed angiotensin-converting enzyme/neprilysin) inhibitors.

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