To determine the molecular basis of the GM2 gangliosidosis 0 variant, we constructed a three-dimensional structure of the human beta-hexosaminidase beta-subunit by homology modeling. It is composed of two domains, domains I and II, and has three disulfide bonds. C534 is located on an extra helix in domain II and forms a disulfide bond with C551. The extra helix is structurally located near domain I. C534Y, identified in a patient with the infantile form of the disease, was deduced to cause disruption of the disulfide bond, which results in a large conformational change of the extra helix, stabilizing the two domains. The drastic change in the protein structure results in a deficiency of the mature beta-subunit, and deficient activities of beta-hexosaminidases A (abeta) and B (betabeta), followed by abundant accumulation of GM2 ganglioside in the patient's cells. R505 is located on the eighth helix of domain II. R505Q, found in a patient with the chronic form of the disease, is predicted to influence the surface structure of the beta-subunit, although it does not affect the active site. The amino acid substitution causes a partial processing defect and decreased enzyme activities, which result in moderate accumulation of GM2 ganglioside in the patient's cells. The structural defects well reflect biochemical and phenotypic abnormalities of the disease.