Indication for a new lipolytic enzyme family: isolation and characterization of two esterases from a metagenomic library.

We have isolated several novel esterase genes from a sheep rumen metagenomic library using the activity-based cluster screening approach as a highly efficient screening technology. The two most remarkable esterase genes, designated estGK1 and estZ3, were further examined. Sequence analysis of estGK1 and estZ3 revealed that they encoded proteins covering 322 and 317 amino acids, respectively. Both proteins were biochemically characterized. EstGK1 and EstZ3 have only minor overall sequence similarity to known esterases. We propose that, together with other hypothetical enzymes, they constitute a new family of lipolytic enzymes. EstGK1 harbors the catalytic serine in the conserved pentapeptide GHSQG, which is typical for lipases, whereas EstZ3 and several other hypothetical proteins contain the pentapeptide SHSQG, a new variation of the conserved motif in lipolytic enzyme families.