Candida maltosa JCM1504 can grow well on L-alanine as a sole carbon and nitrogen source. We found that the activities of alanine aminotransferase (AlaAT) and NAD-dependent glutamate dehydrogenase were remarkably induced when glucose-grown cells were transferred to medium containing L-alanine. This suggested that C. maltosa has an inducible L-alanine degradation system including the above two enzymes. To assess whether AlaAT is essential for the first step of L-alanine degradation, we isolated mutant N-07, which was unable to use L-alanine as a nitrogen source, from the wild strain. Mutant N-07 was very similar to the wild strain in terms of growth on pyruvate and on various amino acids other than L-alanine, suggesting that N-07 lacked only L-alanine-assimilating ability. The AlaAT activity in the cell extract of N-07 was very low and was not induced by L-alanine, whereas the NAD-dependent glutamate dehydrogenase activity was the same as that of the wild strain and was inducible. Western blots with antibody raised against purified AlaAT from C. maltosa indicated that no AlaAT protein was expressed in the mutant N-07. The low level of AlaAT activity described above was possibly due to the pyruvate-forming activity of other enzymes under the assay conditions. From these results, we concluded that AlaAT is an indispensable key enzyme for L-alanine assimilation in C. maltosa.