HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Supplemental Research Data All Versions of this Article: ps.062718507v1 16/8/1522    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Braun, O. Articles by Vasák, M. Search for Related Content PubMed PubMed Citation Articles by Braun, O. Articles by Vasák, M. Social Bookmarking                   What's this?

Specific reactions of S-nitrosothiols with cysteine hydrolases: A comparative study between dimethylargininase-1 and CTP synthetase

Oliver Braun, Markus Knipp¹, Serge Chesnov², and Milan Vaák

Department of Biochemistry, University of Zürich, CH-8057 Zürich, Switzerland

(RECEIVED December 12, 2006; FINAL REVISION April 27, 2007; ACCEPTED April 27, 2007)

S-Transnitrosation is an important bioregulatory process whereby NO⁺ equivalents are transferred between S-nitrosothiols and Cys of target proteins. This reaction proceeds through a common intermediate R–S–N(O^–)–S–R' and it has been proposed that products different from S-nitrosothiols may be formed in protein cavities. Recently, we have reported on the formation of such a product, an N-thiosulfoximide, at the active site of the Cys hydrolase dimethylargininase-1 (DDAH-1) upon reaction with S-nitroso-L-homocysteine (HcyNO). Here we have addressed the question of whether this novel product can also be formed with the endogenously occurring S-nitrosothiols S-nitroso-L-cysteine (CysNO) and S-nitrosoglutathione (GSNO). Further, to explore the reason responsible for the unique formation of an N-thiosulfoximide in DDAH-1 we have expanded these studies to cytidine triphosphate synthetase (CTPS), which shows a similar active site architecture. ESI-MS and activity measurements showed that the bulky GSNO does not react with both enzymes. In contrast, S-nitrosylation of the active site Cys occurred in DDAH-1 with CysNO and in CTPS with CysNO and HcyNO. Although kinetic analysis indicated that these compounds act as specific irreversible inhibitors, no N-thiosulfoximide was formed. The reasons likely responsible for the absence of the N-thiosulfoximide formation are discussed using molecular models of DDAH-1 and CTPS. In tissue extracts DDAH was inhibited only by HcyNO, with an IC₅₀ value similar to that of the isolated protein. Biological implications of these studies for the function of both enzymes are discussed.

Keywords: CTP synthetase; dimethylargininase; glutamine amidotransferase; hyperhomocyst(e)inemia; S-nitrosothiols; nitric oxide; S-transnitrosation

² Functional Genomics Center Zürich, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.

Supplemental material: see www.proteinscience.org

Reprint requests to: Milan Vaák, Department of Biochemistry, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland; e-mail: mvasak{at}bioc.uzh.ch; fax: +41-44-635-59-05.

Abbreviations: ADI, arginine deiminase; ADMA, N,N-dimethyl-L-arginine; AT, Arg:Gly amidinotransferase; Cit, L-citrulline; CTPS, cytidine triphosphate synthetase; CysNO, S-nitroso-L-cysteine; DDAH, N,N -dimethyl-L-arginine dimethylaminohydrolase; DTT, 1,4-dithio-D,L-threitol; EcCTPS, Escherichia coli cytidine triphosphate synthetase; ESI, electrospray ionization; GATase, glutamine amidotransferase; glyco-SNAP, N-(-D-glucopyranosyl)-N ²-acetyl-S-nitroso-D,L-penicillaminamide; GSH, glutathione; GSNO, S-nitrosoglutathione; Hcy, L-homocysteine; HcyNO, S-nitroso-L-homocysteine; HEPES, 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid; MMA, N -methyl-L-arginine; NOS, nitric oxide synthase; PAD4, peptidylarginine deiminase 4; PTP-1B, protein tyrosine phosphatase 1B; Q-TOF, quadrupole time-of-flight; RSH, thiol; RSNO, S-nitrosothiol SNAP, S-nitroso-N-acetyl-D,L-penicillamine; TCEP, tris(2-carboxyethyl)phosphine·HCl; TCA, trichloroacetic acid; TES, N-[tris(hydroxymethyl)methyl]-2-aminoethanesulfonic acid.

Article published online ahead of print. Article and publication are at http://www.proteinscience.org/cgi/doi/10.1110/ps.062718507.

CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				L. Hong and W. Fast Inhibition of Human Dimethylarginine Dimethylaminohydrolase-1 by S-Nitroso-L-homocysteine and Hydrogen Peroxide: ANALYSIS, QUANTIFICATION, AND IMPLICATIONS FOR HYPERHOMOCYSTEINEMIA J. Biol. Chem., November 30, 2007; 282(48): 34684 - 34692. [Abstract] [Full Text] [PDF]