The acyl transferase from Mycobacterium smegmatis (MsAcT) catalyses transesterification reactions in aqueous media because of its hydrophobic active site. Aliphatic cyanohydrin and alkyne esters can be synthesised in water with excellent and strikingly opposite enantioselectivity [(R);E>37 and (S);E>100, respectively]. When using this enzyme, the undesired hydrolysis of the acyl donor is an important factor to take into account. Finally, the choice of acyl donor can significantly influence the obtained enantiomeric excesses.

Acyltransferase from Mycobacterium smegmatis (MsAcT) immobilised in continuous-flow microchannel (30-50 ?m dia.) reactors with hierarchical pore structure (4 cm3/g total pore volume) enabled quantitative, full and rapid transesterification of neopentylglycol with ethyl acetate in a biphasic 50/50 % system in less than one minute. MsAcT was attached either covalently via amino groups or by a specific His-tag-mediated adsorption on Ni or Co sites. Both methods gave similar results for enzyme loading (ca.3 mg/g of carrier, 60-70 % immobilisation yield) and specific activity. The experiments revealed that the rate of monoester formation in the microreactor was exceedingly fast compared to that of diester synthesis and also the native enzyme behaviour in batch reactor. The studies show that the course of transesterification was fully controlled by biocatalytic properties of MsAcT confined in the mesoporous environment. These findings may be of significant interest from both fundamental and practical perspective.