Functional metagenomics reveals novel β-galactosidases not predictable from gene sequences

A soil metagenomic library carried in pJC8 (an IncP cosmid) was used for functional complementation for {beta}-galactosidase activity in both -Proteobacteria (Sinorhizobium meliloti) and{gamma} -Proteobacteria (Escherichia coli). One {beta}-galactosidase, encoded by overlapping clones selected in both hosts, was identified as a member of glycoside hydrolase family 2. ORFs obviously encoding possible {beta}-galactosidases were not identified in 19 other clones that were only able to complement S. meliloti. Based on low sequence similarity to known glycoside hydrolases but not {beta}-galactosidases, three ORFs were examined further. Biochemical analysis confirmed that all encoded {beta}-galactosidase activity. Bioinformatic and structural modeling implied that Lac161_ORF10 protein represented a novel enzyme family with a five-bladed propeller glycoside hydrolase domain.