Variants of isovaleryl-CoA dehydrogenase (IVDHase, EC 1.3.99.10) in 15 isovaleric acidemia fibroblast lines were analyzed using [35S]methionine labeling, immunoprecipitation with anti-rat IVDHase antiserum, and NaDodSo4/polyacrylamide gel electrophoresis. Five distinct variants of IVDHase were detected. The molecular size of variant 1 (43 kDa) was indistinguishable from that of normal IVDHase (43 kDa), although the activity of this enzyme was as deficient (0-2.2% of normal control) as that of any other variant. It was synthesized as a precursor (45 kDa), which is the case for normal IVDHase. Variant 2 was synthesized as a 42-kDa precursor, but only a small portion of it was processed to the mature variant form (40 kDa). Variant 3 (41 kDa) was synthesized as a 43-kDa precursor. Variant 4 (40 kDa) was synthesized as a 42-kDa precursor that was readily processed to the mature form. In cells with variant 5, no material that crossreacted with the anti-rat IVDHase antibody was detected. These results suggest that variant 1 may be due to a point mutation, while variants 2-4 may be encoded by a different mutant IVDHase allele that causes the premature termination of translation, although other complex mechanisms are possible. A deletion, a nonsense mutation close to the NH2 terminus or an extremely labile mRNA may give rise to variant 5.