The esterase from Alicyclobacillus acidocaldarius as a reporter enzyme and affinity tag for protein biosynthesis

Esterase from thermophilic bacteria Alicyclobacillus acidocaldarius can be produced up to 200 mu g/ml by coupled in vitro transcription/translation system derived from Escherichia coli. The synthesized thermostable enzyme can be determined by photometrical and fluorescent assays at least up to 10(-8) M concentration or by activity staining in the polyacrylamide gels. Enhanced green fluorescence protein-esterase fusion protein was bound to a matrix with immobilized esterase inhibitor and purified by affinity chromatography. Thus, the esterase is suited as a reporter enzyme to monitor the expression of polypeptides coupled to its N-terminus and simultaneously, as a cleavable tag for polypeptide purification. (c) 2005 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.