This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Ikushiro, H.
Right arrow Articles by Hayashi, H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ikushiro, H.
Right arrow Articles by Hayashi, H.

 Previous Article  |  Next Article 

Journal of Bacteriology, August 2007, p. 5749-5761, Vol. 189, No. 15
0021-9193/07/$08.00+0     doi:10.1128/JB.00194-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Molecular Characterization of Membrane-Associated Soluble Serine Palmitoyltransferases from Sphingobacterium multivorum and Bdellovibrio stolpii{triangledown}

Hiroko Ikushiro,1* Mohammad Mainul Islam,1,{dagger} Hiromasa Tojo,2 and Hideyuki Hayashi1*

Department of Biochemistry, Osaka Medical College, Takatsuki, Osaka 569-8686, Japan,1 Department of Biochemistry and Molecular Biology, Osaka University, Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan2

Received 5 February 2007/ Accepted 10 May 2007

Serine palmitoyltransferase (SPT) is a key enzyme in sphingolipid biosynthesis and catalyzes the decarboxylative condensation of L-serine and palmitoyl coenzyme A (CoA) to form 3-ketodihydrosphingosine (KDS). Eukaryotic SPTs comprise tightly membrane-associated heterodimers belonging to the pyridoxal 5'-phosphate (PLP)-dependent {alpha}-oxamine synthase family. Sphingomonas paucimobilis, a sphingolipid-containing bacterium, contains an abundant water-soluble homodimeric SPT of the same family (H. Ikushiro et al., J. Biol. Chem. 276:18249-18256, 2001). This enzyme is suitable for the detailed mechanistic studies of SPT, although single crystals appropriate for high-resolution crystallography have not yet been obtained. We have now isolated three novel SPT genes from Sphingobacterium multivorum, Sphingobacterium spiritivorum, and Bdellovibrio stolpii, respectively. Each gene product exhibits an ~30% sequence identity to both eukaryotic subunits, and the putative catalytic amino acid residues are conserved. All bacterial SPTs were successfully overproduced in Escherichia coli and purified as water-soluble active homodimers. The spectroscopic properties of the purified SPTs are characteristic of PLP-dependent enzymes. The KDS formation by the bacterial SPTs was confirmed by high-performance liquid chromatography/mass spectrometry. The Sphingobacterium SPTs obeyed normal steady-state ordered Bi-Bi kinetics, while the Bdellovibrio SPT underwent a remarkable substrate inhibition at palmitoyl CoA concentrations higher than 100 µM, as does the eukaryotic enzyme. Immunoelectron microscopy showed that unlike the cytosolic Sphingomonas SPT, S. multivorum and Bdellovibrio SPTs were bound to the inner membrane of cells as peripheral membrane proteins, indicating that these enzymes can be a prokaryotic model mimicking the membrane-associated eukaryotic SPT.

* Corresponding author. Mailing address: Department of Biochemistry, Osaka Medical College, Takatsuki, Osaka 569-8686, Japan. Phone: 81-72-684-7291. Fax: 81-72-684-6516. E-mail for H. Ikushiro: ikushiro{at}art.osaka-med.ac.jp. E-mail for H. Hayashi: hayashi{at}art.osaka-med.ac.jp

{triangledown} Published ahead of print on 8 June 2007.

{dagger} Present address: Department of Biochemistry, Wake Forest University School of Medicine, Winston-Salem, NC 27157.

Journal of Bacteriology, August 2007, p. 5749-5761, Vol. 189, No. 15
0021-9193/07/$08.00+0     doi:10.1128/JB.00194-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Raman, M. C. C., Johnson, K. A., Yard, B. A., Lowther, J., Carter, L. G., Naismith, J. H., Campopiano, D. J. (2009). The External Aldimine Form of Serine Palmitoyltransferase: STRUCTURAL, KINETIC, AND SPECTROSCOPIC ANALYSIS OF THE WILD-TYPE ENZYME AND HSAN1 MUTANT MIMICS. J. Biol. Chem. 284: 17328-17339 [Abstract] [Full Text]  
  • Pruett, S. T., Bushnev, A., Hagedorn, K., Adiga, M., Haynes, C. A., Sullards, M. C., Liotta, D. C., Merrill, A. H. Jr. (2008). Thematic Review Series: Sphingolipids. Biodiversity of sphingoid bases ("sphingosines") and related amino alcohols. J. Lipid Res. 49: 1621-1639 [Abstract] [Full Text]  
  • Ikushiro, H., Fujii, S., Shiraiwa, Y., Hayashi, H. (2008). Acceleration of the Substrate C{alpha} Deprotonation by an Analogue of the Second Substrate Palmitoyl-CoA in Serine Palmitoyltransferase. J. Biol. Chem. 283: 7542-7553 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»