Activation of soluble guanylate cyclase in the presence of purified human aldehyde dehydrogenases

BMC Pharmacology, Jul 2007

Matteo Beretta, Alexander Kollau, Michael Russwurm, Doris Koesling, Wing Keung, Kurt Schmidt, Bernd Mayer

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Activation of soluble guanylate cyclase in the presence of purified human aldehyde dehydrogenases

Matteo Beretta 2 Alexander Kollau 2 Michael Russwurm 1 Doris Koesling 1 Wing Ming Keung 0 Kurt Schmidt 2 Bernd Mayer 2 0 Department of Pathology, Harvard Medical School , Boston, 02115, MA , USA 1 Department of Pharmacology and Toxicology, Ruhr-Universitat Bochum , 44780 Bochum , Germany 2 Department of Pharmacology and Toxicology, Karl-Franzens-Universitat Graz , 8010 Graz , Austria <supplement><title><p>3<sup>rd</sup>InternationalConferenceoncGMPGenerators,EfectorsandTherapeuticImplications</p></title><note>Me tingabstracts-AsinglePDFcontainingal abstractsinthisSup lementisavailable<ahref="htp:/www.biomedcentral.com/content/files/pdf/1471-2210-7-S1-ful.pdf">here</a>.</note></sup lement> This abstract is available from: http://www.biomedcentral.com/1471-2210/7/S1/P7 - Mitochondrial aldehyde dehydrogenase (ALDH2) was reported to catalyze bioactivation of nitroglycerin (GTN), resulting in the formation of a nitric oxide (NO) like species activating soluble guanylate cyclase (sGC). However, the reaction product is thought to be nitrite, which does not activate sGC, so the link between GTN metabolism and sGC activation is not clear yet. Since we found no link between ALDH2-catalyzed GTN metabolism and mitochondrial nitrite reduction (Kollau A., Beretta M. & Mayer B.; unpublished), we investigated the possibility that an activator of sGC is produced directly in the course of ALDH2 reaction by co-incubation of purified human ALDH2 with purified sGC in the presence of GTN. In the absence of ALDH2, GTN had no effect on sGC activity (0.28 0.07 mol/mg/min). In the presence of 25 g of ALDH2 there was a biphasic stimulation of cGMP formation with an EC50 (half-maximally effective concentration) of ~1 M GTN and a maximum at 10 M GTN (1.66 0.22 mol/mg/min). The effect of ALDH2 was dependent on the protein concentration, with a linear increase from 1 to 100 g (2.10 0.23 mol/ mg/min) and saturation between 100 and 250 g of ALDH2. ALDH2-dependent sGC activation was inhibited by 1 mM chloral hydrate (41.5% of control) and by 100 M daidzin (18.9% of control). The effect of ALDH2 on cGMP formation was almost completely inhibited by the NO scavenger oxy-haemoglobin, the superoxide generator flavin adenine dinucleotide, and the heme site sGC inhibitor ODQ (0.1 mM, each). The cytosolic ALDH isoform (ALDH1) also metabolized GTN to 1,2- and 1,3-GDN (dinitroglycerin) and triggered sGC activation. In line with the about 1,000-fold lower substrate affinity of ALDH1, significantly higher GTN concentrations were required for sGC activation. There was a linear increase in cGMP formation by sGC co-incubated with 50 g of ALDH1 and 10 M to 1 mM GTN. At the highest GTN concentration tested (1 mM), sGC activity was 4.25 0.15 mol/mg/min. As observed with ALDH2mediated sGC activation, the response to ALDH1 was dependent on the protein concentration with a linear increase from 5 to 100 g ALDH1. The effect of ALDH1 on cGMP formation was inhibited by 1 mM chloral hydrate (57% of control) but not by 100 M daidzin. These results suggest that both ALDH1 and ALDH2 catalyze GTN bioactivation by direct formation of NO or a NO-like activator of sGC. (...truncated)


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Matteo Beretta, Alexander Kollau, Michael Russwurm, Doris Koesling, Wing Keung, Kurt Schmidt, Bernd Mayer. Activation of soluble guanylate cyclase in the presence of purified human aldehyde dehydrogenases, BMC Pharmacology, 2007, pp. P7, 7,