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. 1999 Nov;65(11):4873-80.
doi: 10.1128/AEM.65.11.4873-4880.1999.

Genetic characterization of the major lactococcal aromatic aminotransferase and its involvement in conversion of amino acids to aroma compounds

L Rijnen  1 , S Bonneau, M Yvon
Affiliations

Genetic characterization of the major lactococcal aromatic aminotransferase and its involvement in conversion of amino acids to aroma compounds

L Rijnen et al. Appl Environ Microbiol. 1999 Nov.

Abstract

In lactococci, transamination is the first step of the enzymatic conversion of aromatic and branched-chain amino acids to aroma compounds. In previous work we purified and biochemically characterized the major aromatic aminotransferase (AraT) of a Lactococcus lactis subsp. cremoris strain. Here we characterized the corresponding gene and evaluated the role of AraT in the biosynthesis of amino acids and in the conversion of amino acids to aroma compounds. Amino acid sequence homologies with other aminotransferases showed that the enzyme belongs to a new subclass of the aminotransferase I subfamily gamma; AraT is the best-characterized representative of this new aromatic-amino-acid-specific subclass. We demonstrated that AraT plays a major role in the conversion of aromatic amino acids to aroma compounds, since gene inactivation almost completely prevented the degradation of these amino acids. It is also highly involved in methionine and leucine conversion. AraT also has a major physiological role in the biosynthesis of phenylalanine and tyrosine, since gene inactivation weakly slowed down growth on medium without phenylalanine and highly affected growth on every medium without tyrosine. However, another biosynthesis aromatic aminotransferase is induced in the absence of phenylalanine in the culture medium.

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Figures

FIG. 1
FIG. 1
Schematic representation of the constructions of mutants with the luxAB genes as reporters under the control of the araT promoter. (A) Chromosome region containing araT in the wild-type strain; (B) construction with araT intact; (C) construction with araT interrupted. formula image, the region upstream of araT with two potential promoters and a potential terminator just upstream.
FIG. 2
FIG. 2
Alignment of the fingerprints of subfamilies α, β, and γ of aminotransferase family I (according to Jensen and Gu [22]) with the amino acid sequence of AraT. The fingerprint residues conserved in AraT are bold and underlined. Asterisks indicate conserved residues of the subfamily α hinge region.
FIG. 3
FIG. 3
Reverse-phase HPLC separation and identification of [3H]phenylalanine metabolites produced by incubation for 40 h of resting wild-type cells (WT) and araT mutant cells (ArAT−) in a reaction mixture containing α-ketoglutarate under the conditions described in Materials and Methods. C, refers to the reaction mixture with the cells of the wild type before incubation (time zero) (control).
FIG. 4
FIG. 4
Amino acid degradation by wild-type cells (A) and araT mutant cells (B) after incubation for 10 h (black bars), 20 h (hatched bars), and 40 h (white bars) of resting cells in reaction medium containing radiolabeled amino acids as a tracer and α-ketoglutarate, under conditions described in Materials and Methods. Abbreviations: phe, phenylalanine; tyr, tyrosine; trp, tryptophan; leu, leucine; ile, isoleucine; val, valine; met, methionine. Error bars indicate standard deviations of triplicate determinations.
FIG. 5
FIG. 5
Phenylalanine (phe) and phenylpyruvate (ppy) aminotransferase activities of wild-type cells and araT mutant cells grown in CDM and CDM without phenylalanine (CDM-phe). Aminotransferase activities are expressed as glutamate (Glu) or α-ketoglutarate (KG) produced from α-ketoglutarate or glutamate. Error bars indicate standard deviations of triplicate determinations.
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References

    1. Alting A C, Engels W J M, van Schalkwijk S, Exterkate F A. Purification and characterization of cystathionine β-lyase from Lactococcus lactis subsp. cremoris B78 and its possible role in flavor development in cheese. Appl Environ Microbiol. 1995;61:4037–4042. - PMC - PubMed
    1. Altschul S F, Madden T L, Schaffer A A, Zang J, Miller W, Lipman D J. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997;25:3389–3402. - PMC - PubMed
    1. Bardowski J, Ehrlich S D, Chopin A. Tryptophan biosynthesis genes in Lactococcus lactis subsp. lactis. J Bacteriol. 1992;174:6563–6570. - PMC - PubMed
    1. Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72:248–254. - PubMed
    1. Bruinenberg P G, de Roo G, Limsowtin G K V. Purification and characterization of cystathionine γ-lyase from Lactococcus lactis subsp. cremoris SK11: possible role in flavor compound formation during cheese maturation. Appl Environ Microbiol. 1997;63:561–566. - PMC - PubMed

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