The Plethora of Angiotensin-Converting Enzyme-Processed Peptides in Mouse Plasma

Abstract

Angiotensin-converting enzyme (ACE) converts angiotensin I into the potent vasoconstrictor angiotensin II, which regulates blood pressure. However, ACE activity is also essential for other physiological functions, presumably through processing of peptides unrelated to angiotensin. The goal of this study was to identify novel natural substrates and products of ACE through a series of mass-spectrometric experiments. This included comparing the ACE-treated and untreated plasma peptidomes of ACE-knockout (KO) mice, validation with select synthetic peptides, and a quantitative in vivo study of ACE substrates in mice with distinct genetic ACE backgrounds. In total, 244 natural peptides were identified ex vivo as possible substrates or products of ACE, demonstrating high promiscuity of the enzyme. ACE prefers to cleave substrates with Phe or Leu at the C-terminal P2′ position and Gly in the P6 position. Pro in P1′ and Iso in P1 are typical residues in peptides that ACE does not cleave. Several of the novel ACE substrates are known to have biological activities, including a fragment of complement C3, the spasmogenic C3f, which was processed by ACE ex vivo and in vitro. Analyses with N-domain-inactive (NKO) ACE allowed clarification of domain selectivity toward substrates. The in vivo ACE-substrate concentrations in WT, transgenic ACE-KO, NKO, and CKO mice correspond well with the in vitro observations in that higher levels of the ACE substrates were observed when the processing domain was knocked out. This study highlights the vast extent of ACE promiscuity and provides a valuable platform for further investigations of ACE functionality.

Renin‐angiotensin aldosterone profile before and after angiotensin‐converting enzyme‐inhibitor administration in dogs with angiotensin‐converting enzyme gene polymorphism

“…The finding of high aldosterone in conjunction with suppressed angiotensin II formation in some dogs in this study suggests that ABT is occurring independent of angiotensin II. A multitude of peptides serve as substrates for ACE, and the accumulation of these peptides as a result of ACE‐inhibition could be contributing to the formation of aldosterone through other pathways . These findings are important because they demonstrate the efficacy of enalapril in suppressing angiotensin II formation, but also show ABT and indicate that multimodal RAAS suppression may be indicated to address modulation of other pathways by ACE‐inhibitors.…”

“…A multitude of peptides serve as substrates for ACE, and the accumulation of these peptides as a result of ACE-inhibition could be contributing to the formation of aldosterone through other pathways. 23 and inhibition of angiotensin II formation or binding (ACE-inhibitors or angiotensin receptor blockers, respectively) is often necessary to provide comprehensive RAAS inhibition. 18,21,24 Additional study is required to elucidate other pathways that may be impacted by RAAS inhibitors.…”

ACE2 and ACE: structure-based insights into mechanism, regulation and receptor recognition by SARS-CoV

“…The centrality of sACE in regulating the levels of these diverse peptides has made it an attractive drug target for conditions such as hypertension, kidney disease, cardiovascular disease, fibrosis and Alzheimer’s disease. The full extent of this protein’s influence on human health and disease is, however, still unknown, as was emphasised by a recent study which identified more than 240 ACE-regulated peptides in mouse plasma [ 19 ]. Even the ‘traditional’ blood pressure-regulating function of sACE is more complex than originally thought, with cross-talk occurring between the RAAS, natriuretic peptide system (NPS), kallikrein–kinin system (KKS) and the counterregulatory Mas axis of the RAAS [ 20 ].…”

Angiotensin‐converting enzyme open for business: structural insights into the subdomain dynamics

“…In addition to Ang I, peptides that have been identified as substrates for these and potentially other roles include N-acetyl-Ser-Asp-Lys-Pro (AcSDKP), substance P, enkephalins, luteinising hormone-releasing hormone, kinins, the amyloid-beta peptide (Ab), neurotensin and formyl-Met-Leu-Phe [16][17][18][19][20][21]. A recent study which used mass spectrometry to study ACE activity in mouse plasma identified more than 240 potential substrates or products of ACE, demonstrating the vast promiscuity of this enzyme [22]. Furthermore, ACE has been implicated in the immune response through cleavage of Ang I as well as an unknown substrate [23].…”