HexA genes

Tay-Sachs disease. A member of a family of disorders identified as the Gm2 gangliosidoses. As neural cell membranes are enriched in Gm2 gangliosides, the inability to degrade this class of sphingolipid resnlts in neural cell death. In addition to Tay-Sachs disease the family includes the Sandhoff diseases and the Gm2 activator deficiencies. Tay-Sachs disease resnlts from defects in the HEXA gene encoding the a-subunit of /3-hexosaminidase. 

HEXA deficiency has turned out to be neither genetically nor ethnically homogeneous. A variety of different alterations— mutations—in the HEXA gene have been associated with classic, infantile Tay-Sachs disease. The existence of different... 

More important for this discussion, abnormalities of the responsible HEXA gene have now been associated with a dozen or more clinically distinct patterns (Table 1), including schizophrenia and including people with no clinically significant disability. Put technically, adult onset Tay-Sachs disease is clinically pleomorphic. A steady stream of reports continues to appear describing variant neurological abnormalities in people with adult onset Tay-Sachs disease. ... 

Hexosaminidase A, the enzyme defective in Tay-Sachs disease, is actually composed of two subunits, an a and a 3 chain. The exact stoichiometry of the active enzyme is unknown, but it may be aiPi- The a subunit is coded for by the HexA gene, whereas the p subunit is coded for by the HexB gene. In Tay-Sachs disease, the a subunit is defective, and hexosaminidase A activity is lost. However, the p subunit can form active tetramers in the absence of the a subunit, and this activity, named hexosaminidase B, which cleaves the glycolipid globoside, retains activity in children with Tay-Sachs disease. Thus, children with Tay-Sachs disease accumulate the ganglioside GM2, but not globoside (Fig. 30.17). 

Yamanaka, S. Johnson, M.D. Gringerg, A. Westphal, H Crawley, J.N. Taniike, M. Suzuki, K. Proia, R.L. Targeted Disruption of the Hexa Gene Results in Mice with Biochemical and Pathologic Features of Tay-Sachs Disease. Proc. Natl Acad. Sci. USA, 1994, 91, 9975-9979. 

E. coli strain BL21(DE3) (F ompT ksdSafra ms ) gal dcm) (Novagen, Madison, WI, USA) was used as a host for recombinant OPH expression. Recombinant plasmids pTOH and pEOH that contain OPH gene fused with hexa-histidine affinity tag under trc and T7 promoter, respectively, as control vectors and pTTOH and pETOH that contain OPH gene fused with Tat signal sequence and hexa-histidine affinity tag under trc and T7 promoter, respectively, were used (Fig. 1). 

Fig.l. Gene maps of recombinant plasmids pTOH, pTTOH, pEOH, and pETOH. Abbreviations Pnc, trc promoter Px7, T7 and lac hybrid promoter Tat, twin-arginme TorA signal sequence of TMAO reductase OPH, organophosphoms hydrolase gene Hisg, hexa-histidine affinity tag. 

His tag. The ability of the imidazole moiety of the histidine residue to bind divalent metal ions such as nickel, iron, and cobalt can be used to purify histidine-containing proteins on a column which has such divalent metals bound to it. To streamline the purification procedure of any desired protein, such histidines are deliberately added to a protein of choice by cloning a four- to sixfold CAG repeat sequence into the expression construct upstream of the gene of interest so as to express an N-terminal or C-terminal tetra- or hexa-His tag. From experience, especially in E. coli, C-terminal tagging often yields superior results to N-termi-... 

Tay-Sachs disease is caused by the mutation of the alpha subunit of hexosaminidase A gene (HEXA). Deficitated hexosaminidases A and B produce 3 distinct clinical forms of ganglioside GM2 storage disease-Tay-Sachs disease, Sandhoff disease, and juvenile GM2-gangliosidosis. Hexosaminidase-A has a structure comprised of alpha-beta subunits and Tay-Sachs disease is the alpha-minus mutation, whereas Sandhoff disease is a beta-minus mutation (Beutler and Kuhl, 1975 Beutler et al., 1975). Subunit alpha is mapped to chromosome 15 (and beta to chromosome 5). Different levels of residual activities are correlated with the age of clinical onset Tay-Sachs disease, 0.1% of normal hexosaminidase late infantile,... 

CHO AUXBl mutants transfected with the E. coli folylpolyglutamate S3mthetase gene (AUX-co/i) express the E. coli protein in the cytosol and metabolize folates primarily to triglutamates rather than the hexa- and... 

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