Three Neocallimastix patriciarum esterases associated with the degradation of complex polysaccharides are members of a new family of hydrolases

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Abstract
Summary: Acetylesterase and cinnamoyl ester hydrolase activities were demonstrated in culture supernatant of the anaerobic ruminal fungusNeocallimastix patriciarum.A cDNA expression library fromN. patriciarumwas screened for esterases using β-naphthyl acetate and a model cinnamoyl ester compound. cDNA clones representing four different esterase genes (bnaA-D) were isolated. None of the enzymes had cinnamoyl ester hydrolase activity, but two of the enzymes (BnaA and BnaC) had acetylxylan esterase activity.bnaA, bnaBandbnaCencode proteins with several distinct domains. Carboxy-terminal repeats in BnaA and BnaC are homologous to protein-docking domains in other enzymes fromNeocallimastixspecies and another anaerobic fungue, aPiromycessp. The catalytic domains of BnaB and BnaC are members of a recently described family of Ser/His active site hydrolases [Upton, C. & Buckley, J. T. (1995).Trends Biochem Sci20, 178-179]. BnaB exhibits 40% amino acid identity to a domain of unknown function in the CeIE cellulase fromClostridium thermocellumand BnaC exhibits 52% amino acid identity to a domain of unknown function in the XynB xylanase fromRuminococcus flavefaciens.BnaA, whilst exhibiting less than 10% overall amino acid identity to BnaB or BnaC, or to any other known protein, appears to be a member of the same family of hydrolases, having the three universally conserved amino acid sequence motifs. Several other previously described esterases are also shown to be members of this family, including a rhamnogalacturonan acetylesterase fromAspergillus aculeatus.However, none of the other previously described enzymes with acetylxylan esterase activity are members of this family of hydrolases.