Site directed mutagenesis of recombinant pig liver esterase yields mutants with altered enantioselectivity

✍ Scribed by Anna Musidlowska-Persson; Uwe T. Bornscheuer

• Book ID: 104359780
• Publisher: Elsevier Science
• Year: 2003
• Tongue: English
• Weight: 127 KB
• Volume: 14
• Category: Article
• ISSN: 0957-4166
• DOI: 10.1016/s0957-4166(03)00205-2

✦ Synopsis

The enantioselectivity of nine variants of recombinant pig liver esterase (rPLE) including recombinant porcine intestinal carboxyl esterase (rPICE) obtained by site-directed mutagenesis and heterologous expression in Pichia pastoris was investigated. A comparison of these mutants with rPLE revealed significant differences in the kinetic resolution of a series of acetates of secondary alcohols. A six-fold increase in enantioselectivity (E=46) compared to rPLE (E=8) was observed in the hydrolysis of (RS)-1-phenyl-1-ethyl acetate using a variant containing a single mutation (glutamic acid exchanged to glycine at position 77, E77G). No clear correlation between the number and type of amino acid substitutions was found, however a tendency was observed, that at high mutation levels enantioselectivity decreases, but at the same time a switch in enantiopreference occurred as observed for the resolution of (RS)-1-phenyl-3-butyl acetate and (RS)-1-phenyl-2-pentyl acetate.