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

J. Biol. Chem., Vol. 281, Issue 36, 26437-26443, September 8, 2006

This Article Full Text Full Text (PDF) All Versions of this Article: 281/36/26437    most recent M606085200v1 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 Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Robertson, A. S. Articles by Lomas, D. A. Search for Related Content PubMed PubMed Citation Articles by Robertson, A. S. Articles by Lomas, D. A. Social Bookmarking                      What's this?

Inhibitory Activity of the Drosophila melanogaster Serpin Necrotic Is Dependent on Lysine Residues in the D-helix^*

Andrew S. Robertson¹, Didier Belorgey¹, David Gubb^¶, Timothy R. Dafforn^||, and David A. Lomas²

From the Department of Genetics, University of Cambridge, Cambridge CB2 3EH, United Kingdom, the Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Cambridge CB2 2XY, United Kingdom, ^||Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom, and ^¶CIC bioGUNE, Bizkaia Technology Park, 48160 Derio, Spain

Necrotic is a member of the serine protease inhibitor or serpin superfamily. It is a potent inhibitor of elastase and chymotrypsin type proteases and is responsible for regulating the anti-fungal response in Drosophila melanogaster. Necrotic contains three basic lysine residues within the D-helix that are homologous to those found in the heparin-binding domain of antithrombin and heparin co-factor II. We show here that substitution of all three lysine residues for glutamines caused cellular necrosis and premature death in Drosophila in keeping with a loss of function phenotype. The lysine to glutamine substitutions had no effect on the overall structure of recombinant Necrotic protein but abolished the formation of stable complexes with target proteases. Individual substitutions with either glutamine or alanine demonstrated that lysine 68 was the most critical residue for inhibitory activity. Despite the homology to other serpins, Necrotic did not bind, nor was it activated by sulfated glycans. These data demonstrate a critical role for basic residues within the D-helix (and lysine 68 in particular) in the inhibitory mechanism of the serpin Necrotic.

Received for publication, June 26, 2006

^* This work was supported by a Gates Scholarship (to A. S. R.), the Medical Research Council (United Kingdom), the Wellcome Trust, Papworth NHS Trust, Programa Nacional de Biologica Fundemental (Ministerio de Educación y Ciencias, Spain), the Department of Industry, Tourism and Trade of the Government of the Autonomous Community of the Basque Country (Etortek Research Programs 2005/2006), and the Innovation Technology Department of Bizkaia County. 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.

¹ These authors contributed equally to this work.

² To whom correspondence should be addressed: Dept. of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Wellcome Trust/MRC Bldg., Hills Rd., Cambridge CB2 2XY, UK. Tel.: 44-1223-762818; Fax: 44-1223-336827; E-mail: dal16{at}cam.ac.uk.

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

This article has been cited by other articles:

				A. Mulenga, R. Khumthong, and M. A. Blandon Molecular and expression analysis of a family of the Amblyomma americanum tick Lospins J. Exp. Biol., September 15, 2007; 210(18): 3188 - 3198. [Abstract] [Full Text] [PDF]