| Abstract: | Bj-BPP-10c is a bioactive proline-rich decapeptide, part of the C-type natriuretic peptide precursor, expressed in the brain and in the venom gland of Bothrops jararaca. We recently showed that Bj-BPP-10c displays a strong, sustained anti-hypertensive effect in spontaneous hypertensive rats (SHR), without causing any effect in normotensive rats, by a pharmacological effect independent of angiotensin-converting enzyme inhibition. Therefore, we hypothesized that another mechanism should be involved in the peptide activity. Here we used affinity chromatography to search for kidney cytosolic proteins with affinity for Bj-BPP-10c and demonstrate that argininosuccinate synthetase (AsS) is the major protein binding to the peptide. More importantly, this interaction activates the catalytic activity of AsS in a dose-de pend ent manner. AsS is recognized as an important player of the citrulline-NO cycle that represents a potential limiting step in NO synthesis. Accordingly, the functional interaction of Bj-BPP-10c and AsS was evidenced by the following effects promoted by the peptide: (i) increase of NO metabolite production in human umbilical vein endothelial cell culture and of arginine in human embryonic kidney cells and (ii) increase of arginine plasma concentration in SHR. Moreover, α-methyl-dl-aspartic acid, a specific AsS inhibitor, significantly reduced the anti-hypertensive activity of Bj-BPP-10c in SHR. Taken together, these results suggest that AsS plays a role in the anti-hypertensive action of Bj-BPP-10c. Therefore, we propose the activation of AsS as a new mechanism for the anti-hypertensive effect of Bj-BPP-10c in SHR and AsS as a novel target for the therapy of hypertension-related diseases.Inhibition of somatic angiotensin-I-converting enzyme (sACE)3 is a widely used approach in the treatment of hypertension. The first available competitive inhibitors of sACE were the naturally occurring proline-rich oligopeptides from the venom of Bothrops jararaca. Clinical studies using Bj-BPP-9a, teprotide, the most efficient of these snake venom peptides, demonstrated the potential of sACE inhibitors as anti-hypertensive drugs (1). Highly potent inhibitors of sACE, which can be administered orally, have subsequently been developed. The first of these, captopril, was designed employing a theoretical model of the active site of sACE, based on its presumed similarity to the active site of carboxypeptidase A and also with reference to the C terminus of venom proline-rich peptides, which compete with sACE substrates (2). Since captopril reproduced all known pharmacological effects and sACE-inhibiting features of the proline-rich peptides (3), the interest to deepen the investigation of the biological properties of these naturally occurring sACE inhibitors dropped dramatically. However, we recently showed that the Bj-BPP-10c (4), displays a strong and sustained anti-hypertensive effect in spontaneous hypertensive rats (SHR), independently of the inhibition of sACE (5). This result led us to hypothesize that, besides sACE, another molecule involved in the arterial blood pressure homeostasis could possibly be a target for Bj-BPP-10c. Two reasons prompted us to search the putative target in the kidney: (i) the crucial role played by the kidney in the arterial blood pressure control (6) and (ii) the selective concentration and long lasting permanence of 125I-Bj-BPP-10c in the mouse kidney even when a saturating concentration of captopril was administered with the peptide (7).In the present study, we identified the kidney argininosuccinate synthetase (AsS) as a putative target for Bj-BPP10c, and we show results demonstrating that the anti-hypertensive effect of this peptide in SHR is related to the activation of the arginine production in the kidney and of the citrulline-NO cycle in endothelial cells. |
