Active Granzyme A (GZMA)

Granzyme A (GZMA) is a serine protease chiefly released by cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells, playing essential roles in both adaptive and innate immunity. As a central component of the granule exocytosis pathway, GZMA facilitates the eradication of target cells—such as those infected by viruses, transformed tumors, or harboring intracellular pathogens. In contrast to Granzyme B, which predominantly induces mitochondrial apoptosis, GZMA activates a distinct caspase-independent cell death pathway by cleaving specific intracellular substrates, thereby disrupting vital cellular processes. Beyond its direct cytotoxic function, GZMA also contributes to the modulation of inflammatory responses and immune cell activation. Furthermore, it directly interacts with nucleolin (NCL), cleaving it into specific proteolytic fragments.To detect the activity of recombinant GZMA, a functional ELISA assay was performed to evaluate the interaction between recombinant human GZMA and recombinant mouse NCL.Briefly, biotin-linked GZMA were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100µl were then transferred to NCL-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 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 GZMA and NCL was shown in Figure 1, the EC50 for this effect is 0.185µg/mL.