Characterization of the melA locus for alpha-galactosidase in Lactobacillus plantarum.
Alpha-galactosides are abundant sugars in legumes such as soy. Because of the lack of alpha-galactosidase (alpha-Gal) in the digestive tract, humans are unable to digest these sugars, which consequently induce flatulence. To develop the consumption of the otherwise highly nutritional soy products, the use of exogenous alpha-Gal is promising. In this framework, we characterized the melA gene for alpha-Gal in Lactobacillus plantarum. The melA gene encodes a cytoplasmic 84-kDa protein whose enzymatically active form occurs as oligomers. The melA gene was cloned and expressed in Escherichia coli, yielding an active alpha-Gal. We show that melA is transcribed from its own promoter, yielding a monocistronic mRNA, and that it is regulated at the transcriptional level, i.e., it is induced by melibiose but is not totally repressed by glucose. Posttranscriptional regulation by the carbon source could also occur. Upstream of melA, a putative galactoside transporter, designated RafP, was identified that shows high homology to LacS, the unique transporter for both alpha- and beta-galactosides in Streptococcus thermophilus. rafP is also expressed as a monocistronic mRNA. Downstream of melA, the lacL and lacM genes were identified that encode a heterodimeric beta-galactosidase. A putative galM gene identified in the same cluster suggests the presence of a galactose operon. These results indicate that the genes involved in galactoside catabolism are clustered in L. plantarum ATCC 8014. This first genetic characterization of melA and of its putative associated transporter, rafP, in a lactobacillus opens doors to various applications both in the manufacture of soy-derived products and in probiotic and nutraceutical issues.
Silvestroni
Aurelio
A
Centro de Referencia para Lactobacillos (CERELA-CONICET), Chacabuco 145, 4000 Tucumán, Argentina. INRA-URLGA, Useful Bacterial Surface Proteins, 78352 Jouy-en-Josas, France.
Connes
Cristelle
C
Sesma
Fernando
F
De Giori
Graciela Savoy
GS
Piard
Jean-Christophe
JC
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis; D005697:Galactosides / Q000378:metabolism; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D011887:Raffinose / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D027981:Symporters / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D000519:alpha-Galactosidase / Q000235:genetics* / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D005697:Galactosides; D027981:Symporters; C021548:melibiose permease; D000519:alpha-Galactosidase; D001616:beta-Galactosidase; D011887:Raffinose