Combinatorial formulation of biocatalyst preparations for increased activity in organic solvents: Salt activation of penicillin amidase

✍ Scribed by John P. Lindsay; Douglas S. Clark; Jonathan S. Dordick

• Publisher: John Wiley and Sons
• Year: 2004
• Tongue: English
• Weight: 590 KB
• Volume: 85
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/bit.20002

✦ Synopsis

Abstract

A combinatorial experimental technique was used to identify salts and salt mixtures capable of activating penicillin amidase in organic solvents for the transesterification of phenoxyacetate methyl ester with 1‐propanol. Penicillin amidase was lyophilized in the presence of various chloride and acetate salts within 96‐deep‐well plates and catalytic rates measured to determine lead candidates for highly salt‐activated preparations. The kinetics of the most active formulations were then further evaluated. These studies revealed that a formulation consisting of 98% (w/w) of a 1:1 KAc:CsCl salt mixture, 1% (w/w) enzyme, and 1% (w/w) potassium phosphate buffer was ∼35,000‐fold more active than the salt‐free formulation in hexane, as reflected in values of V~max~/K~m~. This extraordinary activation could be extended to more polar solvents, including tert‐amyl alcohol, and to formulations with lower total salt contents. A correlation was found between the kosmotropic/chaotropic behavior of the salts (as measured by the Jones‐Dole B coefficients) and the observed activation. Strongly chaotropic cations combined with strongly kosmotropic anions yielded the greatest activation, and this is likely due to the influence of the ions on protein–water and protein–salt interactions. © 2004 Wiley Periodicals, Inc.