Investigation Of Clinical Diagnosis And Molecular Mechanism Of Sandhoff Disease

Background Sandhoff disease is a rare autosomal recessive lysosomal storage disorder of sphingolipid metabolism resulting from the deficiency of?-hexosaminidase(Hex).Mutations of the HEXB gene cause Sandhoff disease.Objective To investigate the clinical features and gene mutations in the patients with Sandhoff disease,study the phenotype and genotype correlations and provides new insights into the molecular mechanism of Sandhoff disease.Methods In order to improve the diagnosis and expand the knowledge of the disease,we collected and analyzed relevant data of clinical diagnosis,biochemical investigation,and molecular mutational analysis in a group of Chinese patients with Sandhoff disease.In silico analyses were applied to determine the conservation of the mutation sites and pathogenic effects caused by the mutations.The effect of the mutations on the overall organization of the protein and on its activity was also explored using the crystal structure of Hex ? subunit using the bioinformatic software.Results The patients presented with heterogenous symptoms of neurologic deterioration.Hex A and Hex A&B activity in leukocytes was severely deficient.We identified seven different mutations,including three known mutations: IVS12-26G>A,p.Ile207 Val,p.Thr209 Ile,and four novel mutations: p.Leu223 Pro,p.Tyr463 X,p.Pro468 Profs X62,p.Gly549 Arg.We also detected two different heterozygous mutations c.-122 del C and c.-126C>T in the promoter which were suspected to be deleterious mutations.Pedigree analysis indicated Patient 2 was caused by segmental paternal isodisomy with the gene on chromosome 5.The novel mutations p.Tyr463 X and p.Pro468 Profs X62 generated premature stop codon,caused the structure of the enzyme protein incomplete and influenced the function of the enzyme.In regard to the other two novel mutations p.Leu223 Pro and p.Gly549 Arg,conservational analysis showed that the mutation sites were highly conserved among all tested species;functional analysis in sillico predicated that the mutations may be a damaging mutation.Protein crystal structural analysis indicated the mutation p.Thr209 Ile caused the elimination of the hydrogen bonds between residue 209 and both of Trp236 and Ile238,thus changing the tertiary structure of the protein and further impairing the protein function;the mutation p.Gly549 Arg affected the dimerization of Hex ?subunit,thus altering the quarternary structure of the protein and further damaging the protein function;the mutations p.Ile207 Val and p.Leu223 Pro had a few influences on the protein structure and were suggested to still affect the protein function to some extent because they were located at the active center of the enzyme;the mutations p.Tyr463 X and p.Pro468 Profs X62 leaded the elimination of the important structure of the protein,thus destroying the structure and function of the protein.Conclusions Sandhoff disease presented with neurologic deterioration.Hex A and Hex A&B activities in leukocytes were severely deficient.Seven different mutations were identified,including three known mutations and four novel mutations.Bioinformatic analyses suggested the four novel mutations were damaging.Our study expands the spectrum of genotype of Sandhoff disease in China,studies the phenotype and genotype correlations,provides new insights into the molecular mechanism of Sandhoff disease and helps to the diagnosis and treatment of this disease.