MISC

2004年1月

Profiling and comprehensive expression analysis of ABC transporter solute-binding proteins of Bacillus subtilis membrane based on a proteomic approach

ELECTROPHORESIS
  • K Bunai
  • ,
  • M Ariga
  • ,
  • T Inoue
  • ,
  • M Nozaki
  • ,
  • S Ogane
  • ,
  • H Kakeshita
  • ,
  • T Nemoto
  • ,
  • H Nakanishi
  • ,
  • K Yamane

25
1
開始ページ
141
終了ページ
155
記述言語
英語
掲載種別
DOI
10.1002/elps.200305676
出版者・発行元
WILEY-V C H VERLAG GMBH

We analyzed ABC transporter solute-binding proteins (SBPs) of the Bacillus subtilis membrane using a proteomic approach. We prepared a washed cell membrane fraction that was insoluble in 134 mm nondetergent sulfobetaine and then extracted proteins using mixtures of detergents in a stepwise manner. The membrane proteins were resolved by three two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) or two one-dimensional (1-D) PAGE procedures, electroblotted, and digested in the presence of 5% or 80% acetonitrile. Thereafter, matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF MS) identified 637 proteins corresponding to 15.9% of the total cellular proteins. We predicted that among these, 256 were membrane proteins, 101 were lipoproteins or secretory proteins and 280 were soluble proteins containing peripheral proteins that function in both the cytoplasm and the cell membrane such as SecA and FtsY. Among the 637 proteins, we identified 30 SBPs among 38 importers predicted by a bioinformatic search of the genome. We confirmed expression of the genes for the 30 SBPs using DNA microarray analysis. We compared the 2-D gel separation profiles of submembrane fractions solubilized by 1 % n-dodecyl-beta-D-maltoside from cells cultured on Luria Bertani (LB), S7, and S7 medium without glutamate as well as DNA microarray data on LB and S7. The results suggested that YcdH, YtmK and YurO are binding proteins for Mn++, glutamate and glucose, respectively, and that YqiX and YxeM are binding proteins for amino acids (tryptophan in S7 medium).

リンク情報
DOI
https://doi.org/10.1002/elps.200305676
Web of Science
https://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcAuth=JSTA_CEL&SrcApp=J_Gate_JST&DestLinkType=FullRecord&KeyUT=WOS:000188417100019&DestApp=WOS_CPL
ID情報
  • DOI : 10.1002/elps.200305676
  • ISSN : 0173-0835
  • Web of Science ID : WOS:000188417100019

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