Expansion of application of biocatalysts by introduction of organic solvents and non-natural substrates

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It is important to draw out the latent abilities and expand the use of biocatalysts by introducing unusual conditions such as organic solvents and non-natural substrates. We found in this study that halophenylalanines served as non-natural substrates of thermolysin, and the enantioselective synthesis of non-natural peptides containing halophenylalanines was successfully carried out. These non-natural poptides are expected to have novel physiological activities. Furthermore, we tried the bioconversion of organosilicon compounds which play important roles in organic synthesis, and succeeded in the optical resolution of primary alcohols containing stereogenic silicon atom. It was also demonstrated that the dehydrogenation of primary beta-hydroxysilanes proceeded enantioselectively with only alcohol dehydrogenase (HLADH) and a catalytic amount of NAD^+ due to in-situ NAD^+ regeneration. That is, beta-hydroxysilanes were dehydrogenated by HLADH to beta-carbonylsilanes, which were spontaneously degraded into aldehydes, which served substrates for HLADH to regenerate NAD^+. These results indicated that the application of specific features of organosilicon compounds enabled the construction of the efficient bioconversion systems, being impossible with conventional substrates. In addition, it was found that the use of non-natural acid anhydrides as acyl donors in fed-batch system could avoid the accumulation of water which is a big problem of hydrolasecatalyzed esterification. We were also successful in constructing a novel non-support bioreactor needing no conventional immobilization by introducing organic solvent as unusual condition because biocatalysts are not soluble in organic solvents.