Biotransformations of various functionalized racemic nitriles catalyzed by Rhodococcus erythropolis AJ270, a nitrile hydratase/amidase-containing microbial whole-cell catalyst, were studied. While the nitrile hydratase exhibits high catalytic efficiency but very low enantioselectivity against almost all nitrile substrates examined, the amidase is very sensitive toward the structure of the amides. The release of the steric crowdedness around the stereocenter of the substrates and the introduction of an unsaturated carbon-carbon bond into the substrates led to the significant acceleration of the reaction rate and the dramatic enhancement of the enantioselectivity. Nitrile biotransformations provide a unique and high-yielding synthetic route to highly enantiopure carboxylic acids and amides functionalized with an allyl, propargyl, allenyl, or vinyl group. The synthetic applications have been demonstrated by the synthesis of enantiopure heterocyclic compounds including iodoenol gamma-lactone, gamma-lactam, and 3-allyl-1-phenyl-3,4-dihydro-1H-quinolin-2-one derivatives.