Ding_2020_Int.J.Biol.Macromol_148_333

Deacetyl-7-aminocephalosporanic acid (D-7-ACA) is required for producing of many semisynthetic beta-lactam antibiotics; therefore, enzymes capable of converting 7-aminocephalosporanic acid (7-ACA) to D-7-ACA present a valuable resource to the pharmaceutical industry. In the present study, a putative acetylesterase, EstZY, was identified and characterized from a thermophilic bacterium Alicyclobacillus tengchongensis. Sequence alignment showed that EstZY was an acetylesterase which belonged to carbohydrate esterase family 7 (CE7), with substrate preference for short-chain acyl esters p-NPC(2) to p-NPC(8). Maximum enzyme activity was recorded at pH 9.0 and 50 degreesC, where K(m) and V(max) were calculated as 1.9 +/- 0.23 mM and 258 +/- 18.5 microM min(-)(1), respectively. The residues Ser185, Asp274, and His303 were identified as the putative catalytic triad by homology modelling, site-directed mutagenesis and molecular docking. Moreover, EstZY can remove the acetyl group at C3' position of 7-ACA to form D-7-ACA; this is the first report of a 7-ACA deacetylase from CE7 family in A. tengchongensis and may represent a new enzyme with industrial values.