Figure 1. The binding activity of ELA2 with COL17.
Elastase2 (ELA2), elastase 2 is a serine proteinase in the same family as chymotrypsin and has broad substrate specificity. Secreted by neutrophils and macrophages during inflammation, it destroys bacteria and host tissue. The neutrophil form of elastase is 218 amino acids long, with two asparagine-linked carbohydrate chains (see glycosylation). Neutrophil elastase may play a role in degenerative and inflammatory diseases by its proteolysis of collagen-IV and elastin of the extracellular matrix. This protein degrades the outer membrane protein A (OmpA) of E. coli as well as the virulence factors of such bacteria as Shigella, Salmonella and Yersinia. Besides, Collagen Type XVII (COL17) has been identified as an interactor of ELA2, thus a binding ELISA assay was conducted to detect the interaction of recombinant human COL17 and recombinant human ELA2. Briefly, ELA2 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100uL were then transferred to COL17-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-ELA2 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 ELA2 and COL17 was shown in Figure 1, and this effect was in a dose dependent manner.