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

J. Biol. Chem., Vol. 282, Issue 52, 37325-37340, December 28, 2007

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 282/52/37325    most recent M705541200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map 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 Google Scholar Articles by Thijs, L. Articles by Dewilde, S. PubMed PubMed Citation Articles by Thijs, L. Articles by Dewilde, S.

Characterization of a Globin-coupled Oxygen Sensor with a Gene-regulating Function^*

Liesbet Thijs, Evi Vinck¹, Alessandro Bolli^¶, Florin Trandafir, Xuehua Wan^||, David Hoogewijs^**, Massimiliano Coletta, Angela Fago, Roy E. Weber, Sabine Van Doorslaer, Paolo Ascenzi^¶¶¶, Maqsudul Alam^||, Luc Moens, and Sylvia Dewilde²

From the Departments of Biomedical Sciences and Physics, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, the ^¶Dipartimento di Biologia and Laboratorio Interdipartimentale di Microscopia Elettronica, Università Roma Tre, Viale G. Marconi 446, 00146 Roma, Italy, the ^||Department of Microbiology, University of Hawaii, Honolulu, Hawaii 96822, the ^**Department of Biology and Centre for Molecular Phylogeny and Evolution, Ghent University, K .L. Ledeganckstraat 35, 9000 Ghent, Belgium, the Dipartimento di Medicina Sperimentale e Scienze Biochimiche, Università di Roma Tor Vergata, Via Montpellier 1, 00133 Roma, Italy, Zoophysiology, Department of Biological Sciences, University of Aarhus, DK-8000 Aarhus C, Denmark, and ^¶¶Istituto Nazionale per le Malattie Infettive Lazzaro Spallanzani, Via Portuense 292, 00149 Roma, Italy

Globin-coupled sensors (GCSs) are multiple-domain transducers, consisting of a regulatory globin-like heme-binding domain and a linked transducer domain(s). GCSs have been described in both Archaea and bacteria. They are generally assumed to bind O₂ (and perhaps other gaseous ligands) and to transmit a conformational change signal through the transducer domain in response to fluctuating O₂ levels. In this study, the heme-binding domain, AvGReg178, and the full protein, AvGReg of the Azotobacter vinelandii GCS, were cloned, expressed, and purified. After purification, the heme iron of AvGReg178 was found to bind O₂. This form was stable over many hours. In contrast, the predominant presence of a bis-histidine coordinate heme in ferric AvGReg was revealed. Differences in the heme pocket structure were also observed for the deoxygenated ferrous state of these proteins. The spectra showed that the deoxygenated ferrous derivatives of AvGReg178 and AvGReg are characterized by a penta-coordinate and hexa-coordinate heme iron, respectively. O₂ binding isotherms indicate that AvGReg178 and AvGReg show a high affinity for O₂ with P₅₀ values at 20 °C of 0.04 and 0.15 torr, respectively. Kinetics of CO binding indicate that AvGReg178 carbonylation conforms to a monophasic process, comparable with that of myoglobin, whereas AvGReg carbonylation conforms to a three-phasic reaction, as observed for several proteins with bis-histidine heme iron coordination. Besides sensing ligands, in vitro data suggest that AvGReg(178) may have a role in O₂-mediated NO-detoxification, yielding metAvGReg(178) and nitrate.

Received for publication, July 6, 2007 , and in revised form, September 27, 2007.

^* This work was supported by the Fund for Scientific Research of Flanders, the Danish Natural Science Research Council, the Novo Nordisk Foundation, and National Science Foundation Grant MCB0446431. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental text, Tables S1–S2 and Figs. S1–S5.

¹ Research assistant of the Fund for Scientific Research of Flanders.

² Postdoctoral fellow of the Fund of Scientific Research Flanders. To whom correspondence should be addressed: Dept. of Biomedical Sciences, University of Antwerp, Campus Drie Eiken, Universiteitsplein 1, B-2610 Antwerp, Belgium. Tel.: 32-3-8202392; Fax: 32-3-8202339; E-mail: sylvia.dewilde{at}ua.ac.be.