NAD⁺-dependent glycerol 3-phosphate dehydrogenase (GPD) are rate limiting for glycerol production in Saccharomyces cerevisiae. Recently, the gene CgGPD encoding glycerol 3-phosphate dehydrogenase homologous to GPD genes in other yeasts was cloned from Candida glycerinogenes WL2002-5, an excellent industrial glycerol producer. However, the knowledge about CgGPD expression regulation, especially difference with GPD1 and GPD2 from S. cerevisiae is so less. A functional comparison of CgGPD from C. glycerinogenes with GPD1 and GPD2 from S. cerevisiae was undertaken, using S. cerevisiae gpd1/gpd2 and gpd1 osmosensitive mutants as expression systems. The functions of three indicated genes in S. cerevisiae was characterized for osmoregulation under high osmotic stress and redox regulation under anaerobic condition with various transformants. The results showed that gpd1/gpd2 mutants harbouring CgGPD and GPD1 can restore osmotolerance and increase glycerol production ability under hyperosmotic stress but mutant expressed GPD2 can not. When cells were cultured under the anaerobic condition, the growth pattern of mutants harbouring CgGPD and GPD2 are smilar, however the mutant harbouring GPD1 grows slowly and the growth of the controll supresses thoroughly. Furthermore, gpd1/gpd2 mutant employing either CgGPD or GPD2 can increase glycerol production ability and improve GPD specific enzyme activity in anaerobic incubation, but gpd1/gpd2 mutant expressed GPD1 has no the similar results under the same condition. This indicated that CgGPD may involve in both osmoregulation and redox balance and can complement GPD1 and GPD2 in gpd1/gpd2 mutant.