Enzymological and molecular analysis of a novel multiple-substrate aminotransferase from a hyperthermophilic archaeon.

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A hyperthermophilic archaeon, Thermococcus profundus, possessed an amino acid aminotransferase with extremely broad substrate specificity (multiple-substrate aminotransferase ; MsAT). MsAT was composed of two homogeneous subunits of 45,000. The maximal activity was detected at above 80ﾟC and between pH 6.0 and 8.0. Pyridoxal 5'-phosphate was required for the activity. The enzyme catalyzed aminotransfer from L-alanine, L-leucine, L-isoleucine, L-valine, L-phenylalanine, L-tyrosine, L-tryptophan, L-methionine, L-cystein, L-threonine and L-lysine using 2-oxoglutarate as an amino receptor.The gene encoding MsAT was cloned and expressed in Escherichia coil. MsAT revealed only a low degree of sequence identity (less than 25%) to any aminotransferases so far characterized. However, homology search against EMBL database revealed that Pyrococcus horikoshii and Sulfolobus solfataricus possessed an open reading frame homologous to the MsAT gene. MsAT and these homologues formed a novel cluster independent from other aminotransferases on an evolutionary tree based on the amino acid sequences.