Active Hexosaminidase A Alpha (HEXa)

Hexosaminidase A Alpha (HEXa) is a lysosomal enzyme. There are three predominant isoenzymes: hexosaminidase A, B and S. Hexosaminidase A and the cofactor GM2 activator protein catalyze the degradation of the GM2 gangliosides and other molecules containing terminal N-acetyl hexosamines. The enzymes are composed of two alpha and/or beta subunits, which are coded by HEXA and HEXB genes, respectively. Even though the alpha and beta subunits of hexosaminidase A can both cleave GalNAc residues, only the alpha subunit which contains a key residue, Arg-424 is able to hydrolyze GM2 gangliosides. Hexosaminidase A (alpha polypeptide) plays a critical role in the brain and spinal cord (central nervous system). Besides, Hexosaminidase B Beta (HEXB) has been identified as an interactor of HEXA, thus a binding ELISA assay was conducted to detect the interaction of recombinant mouse HEXA and recombinant mouse HEXB. Briefly, HEXA were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100uL were then transferred to HEXB-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-HEXA pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50µL stop solution to the wells and read at 450nm immediately. The binding activity of of HEXA and HEXB was shown in Figure 1, and this effect was in a dose dependent manner.