Use of hydrophobic bacterium Rhodococcus rhodochrous NBRC15564 expressed thermophilic alcohol dehydrogenases as whole-cell catalyst in solvent-free organic media

• The hydrophobic bacteria, Rhodococcus rhodochrous could be well dispersed in organic media.
• R. rhodochrous showed the highest affinity to hydrophobic chemicals.
• R. rhodochrous was used as the biocatalyst in solvent-free organic media.
• The two thermostable alcohol dehydrogenases were expressed in R. rhodochrous.
• Heat-treated R. rhodochrous could convert about 97% of 3.7 M TFAP to TFMBA in 48 h.

The hydrophobic bacterium Rhodococcus rhodochrous NBRC15564 was employed as a whole-cell biocatalyst to examine its potential for bioconversion in solvent-free organic media. The genes encoding two different thermostable alcohol dehydrogenases (ADHTt1 and ADHTt2) of Thermus thermophilus HB27 were expressed in R. rhodochrous cells. To inactivate indigenous mesophilic enzymes in R. rhodochrous, transformant cells were heated at 70 °C for 10 min. Heat-treated hydrophobic wet cells were used for the bioconversion of 2,2,2-trifluoroacetophenone (TFAP) to α-(trifluoromethyl) benzyl alcohol (TFMBA) as a model reaction with ADHTt1. NADH, which was supplied in aqueous solution, was regenerated by converting cyclohexanol to cyclohexanone by ADHTt2. All reactions were performed by suspending heat-treated cells in solvent-free organic media consisting of 3.7 M TFAP and 4.8 M cyclohexanol (1:1, v/v ratio) at 60 °C. When 800 mg heat-treated R. rhodochrous cells were dispersed in 2 mL of solvent-free organic media (400 mg cells/mL), the product concentration reached about 3.6 M TFMBA by 48 h with a total NADH turnover number of approximately 900. The overall productivity was 190 mol TFMBA/kg cells/h.