Cloning and characterization of a beta-galactosidase encoding region in Lactobacillus coryniformis CECT 5711.
A chromosomal DNA fragment of 7.8 kb from Lactobacillus coryniformis CECT 5711 was cloned in Escherichia coli K-12 and was found to express a functional beta-galactosidase. Nucleotide sequence analysis showed that this fragment contained two partially overlapping genes, the lacL (1,881 bp) and the lacM (960 bp), that encode the subunits of a heterodimeric beta-galactosidase, with estimated molecular masses of 72,129 and 35,233 Da, respectively. Other three incomplete open reading frames showing homology to another beta-galactosidase, an alpha-galactosidase, and a galactokinase, respectively, were also found. The L. coryniformis beta-galactosidase was overproduced in E. coli by using an isopropyl-beta-D: -thiogalactopyranoside (IPTG) expression system. Two new proteins with an estimated M (r) s of approximately 72,000 and 35,000 appeared upon induction with IPTG, and extracts of the recombinant E. coli strain showed beta-galactosidase activity.
Corral
J M
JM
Department of Biotechnology, Puleva Biotech, S.A., Camino de Purchil, 66, 18004 Granada, Spain.
Bañuelos
O
O
Adrio
J L
JL
Velasco
J
J
D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D019281:Dimerization; D015723:Gene Library; D005798:Genes, Bacterial; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D001616:beta-Galactosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D004269:DNA, Bacterial; D011994:Recombinant Proteins; D001616:beta-Galactosidase