Pyruvate carboxylase is involved in metabolism of mimosine by Rhizobium sp. strain TAL1145
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ORIGINAL PAPER
Pyruvate carboxylase is involved in metabolism of mimosine by Rhizobium sp. strain TAL1145 Jonathan D. Awaya · Panlada Tittabutr · Qing X. Li · Dulal Borthakur
Received: 7 November 2007 / Revised: 3 March 2008 / Accepted: 29 April 2008 / Published online: 21 May 2008 © Springer-Verlag 2008
Abstract The objective of this study was to determine the role of midK, which encodes a protein similar to pyruvate carboxylase, in mimosine degradation by Rhizobium sp. strain TAL1145. The midK gene is located downstream of midR in the cluster of genes for mimosine degradation in Rhizobium sp. strain TAL1145. The midK mutants of TAL1145 degraded mimosine slower than the wild-type. These mutants could utilize pyruvate as a source of carbon, indicating that there is another pyruvate carboxylase (pyc) gene in TAL1145. Two classes of clones were isolated from the library of TAL1145 by complementing a pyc mutant of Rhizobium etli, one class contained midK, while the other carried pyc. Both midK and pyc of TAL1145 complemented the midK mutant for mimosine degradation, and also the R. etli pyc mutant for pyruvate utilization. The midK-encoded pyruvate carboxylase was required for an eYcient conversion of mimosine into 3-hydroxy-4-pyridone (HP). Keywords Rhizobium · Pyruvate carboxylase · Mimosine degradation
Jonathan D. Awaya and Panlada Tittabutr contributed equally to this work. Communicated by Ursula Priefer. Electronic supplementary material The online version of this article (doi:10.1007/s00203-008-0384-4) contains supplementary material, which is available to authorized users. J. D. Awaya · P. Tittabutr · Q. X. Li · D. Borthakur (&) Department of Molecular Biosciences and Bioengineering, University of Hawaii at Manoa, 1955 East-West Road, Ag. Science 218, Honolulu, HI 96822, USA e-mail: [email protected]
Introduction Rhizobium sp. strain TAL1145 can degrade mimosine, a toxin produced by Leucaena leucocephala (Soedarjo et al. 1994). Mimosine is toxic to animals, plants and microorganisms, because it inhibits cell division by chelating iron from the surrounding (Soedarjo and Borthakur 1998). The mid genes of TAL1145 are involved in conversion of mimosine into 3-hydroxy-4-pyridone (HP), which is then converted to pyruvate, formate and ammonia by enzymes encoded by pydAB genes (Fox and Borthakur 2001; Awaya et al. 2005). The mid genes, which are located within a 10-kb region in the chromosome of Rhizobium sp. strain TAL1145, were cloned in cosmids pUHR181 and pUHR263 (Borthakur et al. 2003). midA, midB and midC encode the proteins of an ABC transporter for uptake of mimosine from the medium by the bacterial cell, while an aminotransferase encoded by midD is required for conversion of mimosine into HP. A lysR-type of regulatory gene, midR, located upstream of midA in the reverse orientation induces the transcription of mid genes in the presence of mimosine. The transconjugants of a mimosine-nondegrading strain (Mid¡) TAL182 containing these mid genes in cosmid pUHR181 converts mimosine into HP. However, the transconju
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