Sulfur-limitation-regulated proteins in Bacillus subtilis: a two-dimensional gel electrophoresis study

Unité de Régulation de l’Expression Génétique, Institut Pasteur, 28 rue du Docteur Roux, 75724 Paris Cedex 15, France 1 Neurobiologie et Diversité Cellulaire, CNRS UMR 7637, Ecole Supérieure de Physique et Chimie Industrielles de la Ville de Paris, 10 rue Vauquelin, 75005 Paris, France 2 INRA, Domaine de Vilvert, 78352 Jouy en Josas, France 3 Author for correspondence: Isabelle Martin-Verstraete. Tel: +33 1 45 68 72 95. Fax: +33 1 45 68 89 48. e-mail: iverstra{at}pasteur.fr Little is known about the genes and enzymes involved in sulfur assimilation in Bacillus subtilis , or about the regulation of their expression or activity. To identify genes regulated by sulfur limitation, the authors used two- dimensional (2D) gel electrophoresis to compare the proteome of a wild-type strain grown with either sulfate or glutathione as sole sulfur source. A total of 15 proteins whose synthesis is modified under these two conditions were identified by matrix-assisted laser desorption/ionization time of flight (MALDI TOF) mass spectrometry. In the presence of sulfate, an increased amount of proteins involved in the metabolism of C 1 units (SerA, GlyA, FolD) and in the biosynthesis of purines (PurQ, Xpt) and pyrimidines (Upp, PyrAA, PyrF) was observed. In the presence of glutathione, the synthesis of two uptake systems (DppE, SsuA), an oxygenase (SsuD), cysteine synthase (CysK) and two proteins of unknown function (YtmI, YurL) was increased. The changes in expression of the corresponding genes, in the presence of sulfate and glutathione, were monitored using slot-blot analyses and lacZ fusions. The ytmI gene is part of a locus of 12 genes which are co-regulated in response to sulfur availability. This putative operon is activated by a LysR-like regulator, YtlI. This is the first regulator involved in the control of expression in response to sulfur availability to be identified in B. subtilis. Keywords: sulfur metabolism, proteome analysis, regulation Abbreviations: 2D, two-dimensional; ABC, ATP-binding cassette; DIG, digoxigenin; MALDI-TOF, matrix-assisted laser desorption/ionization time of flight a Present address: Genopole, Institut Pasteur, 28 rue du Docteur Roux, 75724 Paris Cedex 15, France. c Present address: HKU Pasteur Research Center, Dexter HC Man Building, 8 Sassoon Road, Pokfulam, Hong Kong. b Present address: Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, UK.