Strategic Selection of Hyperthermophilic Esterases for Resolution of 2-Arylpropionic Esters

摘要

Homologues to Carboxylesterase NP and Candida ruosa lipase,used for the chiral separation of racemic mixtures of 2-arylpropionic methyl esters,were identified by BLAST searches of afailable genome sequences for hyperthermophilic microoranisms.Two potential candidates were identified:a putative lysophospholipase from Pyrococcus fureiosus (Pfu-LPL) and a carboxylesterase from Sulfolous solfatarcus P1 (Sso-EST1).Although both enzymes showed hyderolytic preference toward the (S) merhyl ester,only Sso-EST1 yielded highly optically pure(S) naproxen (%ee_p>=90) and was thus further investigated.Changes in pH or reaction time showed littrle improvement in %ee_p or E values wih Sso-EST1.However,the addition of 25% methanol resulted in a 25% increase in E.The effec of vaious cosolventrs on the enantiomeric ratio showed no corelation with the log P or dielectric constrant values of the solvent.Howevere,an inverse relationship between E and the denaturation capacity (DC) of the waer miscible cosolvents was oserved.This was attribited to an increase in enzyme flexibility with increasing solvent DC values leading to a concomitant reduction in the resolving power of Sso-EST1.The results here show that although bioinformation tools can be used to select candidate biocatalysts for chiral resolution of 2-arylpropionic esters,biochemical characterization is needed to definitively determine functional characteristics.