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 over-produced in E. coli by using an isopropyl-beta-D-thiogalacto-pyranoside (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.