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 Atiles, M. W.
Right arrow Articles by Steele, J. L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Atiles, M. W.
Right arrow Articles by Steele, J. L.
Agricola
Right arrow Articles by Atiles, M. W.
Right arrow Articles by Steele, J. L.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, June 2000, p. 2325-2329, Vol. 66, No. 6
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Gene Cloning, Sequencing, and Inactivation of the Branched-Chain Aminotransferase of Lactococcus lactis LM0230

Myrta W. Atiles,1 Edward G. Dudley,1 and James L. Steele2,*

Departments of Bacteriology1 and Food Science,2 University of Wisconsin---Madison, Madison, Wisconsin 53706

Received 7 October 1999/Accepted 10 March 2000

A branched-chain aminotransferase gene (ilvE) from Lactococcus lactis LM0230 was identified on a 9-kb chromosomal insert by complementation in Escherichia coli DL39. Sequencing of a 2.0-kbp fragment resulted in the identification of a 1,023-bp open reading frame that could encode a 340-amino-acid protein. Sequence analysis of the deduced amino acid sequence revealed 62% identity to IlvE of Haemophilus influenzae and high similarity to IlvEs from a variety of organisms found in GenBank classified as class IV aminotransferases. Under logarithmic growth in complex medium, ilvE is transcribed monocistronically as a 1.1-kb transcript. Hydrophobicity plot analysis of the deduced amino acid sequence and the lack of a signal peptide sequence suggest IlvE is a cytosolic protein. A derivative of LM0230 lacking IlvE activity was constructed by gene replacement. Comparison of the IlvE-deficient strain's ability to grow in defined media lacking an amino acid but containing its alpha -keto acid biosynthetic precursor to that of the wild-type strain indicated that IlvE is the only enzyme capable of synthesis of Ile and Val from their biosynthetic precursors. Comparison of the aminotransferase activity of the IlvE mutant to LM0230 revealed that the mutant retained <2, 4.5, 43, 40, and 76% of its aminotransferase activity with Ile, Val, Leu, Met, and Phe, respectively. No difference in growth or acidification rate between LM0230 and the IlvE-deficient strain was observed in milk.

* Corresponding author. Mailing address: Department of Food Science, University of Wisconsin-Madison, Madison, WI 53706. Phone (608) 262-5960. Fax: (608) 262-6872. E-mail: jlsteele{at}facstaff.wisc.edu.

Applied and Environmental Microbiology, June 2000, p. 2325-2329, Vol. 66, No. 6
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Liu, M., Nauta, A., Francke, C., Siezen, R. J. (2008). Comparative Genomics of Enzymes in Flavor-Forming Pathways from Amino Acids in Lactic Acid Bacteria. Appl. Environ. Microbiol. 74: 4590-4600 [Full Text]  
  • Ganesan, B., Dobrowolski, P., Weimer, B. C. (2006). Identification of the Leucine-to-2-Methylbutyric Acid Catabolic Pathway of Lactococcus lactis.. Appl. Environ. Microbiol. 72: 4264-4273 [Abstract] [Full Text]  
  • Ganesan, B., Weimer, B. C. (2004). Role of Aminotransferase IlvE in Production of Branched-Chain Fatty Acids by Lactococcus lactis subsp. lactis. Appl. Environ. Microbiol. 70: 638-641 [Abstract] [Full Text]  
  • Madsen, S. M., Beck, H. C., Ravn, P., Vrang, A., Hansen, A. M., Israelsen, H. (2002). Cloning and Inactivation of a Branched-Chain-Amino-Acid Aminotransferase Gene from Staphylococcus carnosus and Characterization of the Enzyme. Appl. Environ. Microbiol. 68: 4007-4014 [Abstract] [Full Text]  
  • Aubel, D., Germond, J. E., Gilbert, C., Atlan, D. (2002). Isolation of the patC gene encoding the cystathionine {beta}-lyase of Lactobacillus delbrueckii subsp. bulgaricus and molecular analysis of inter-strain variability in enzyme biosynthesis. Microbiology 148: 2029-2036 [Abstract] [Full Text]  
  • Dudley, E. G., Steele, J. L. (2001). Lactococcus lactis LM0230 contains a single aminotransferase involved in aspartate biosynthesis, which is essential for growth in milk. Microbiology 147: 215-224 [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»