The present invention relates to a process of producing isobutanol, including: mixing water, lactate, an enzyme mixture including at least one enzyme, at least one cofactor, and at least one coenzyme, to prepare a reaction mixture; allowing catalytic conversions of lactate in the reaction mixture for a sufficient amount of time to produce isobutanol; and separating the isobutanol from a reactant obtained by the catalytic conversions, in which the conversion of lactate into isobutanol is in association with a NAD+/NADH and/or NADP+/NADPH regenerating system.