Dibasic endoprotease (also known as Kex2 protease, YSCF protease, EC 3.4.21.61) is responsible for processing the precursors of killer toxin and alpha-factor in yeast. It is a calcium ion-dependent serine protease that specifically recognizes and cleaves peptide bonds at the carboxyl terminus of dibasic amino acid pairs such as Arg-Arg and Lys-Arg.

Kex2 protease cleaves the carboxyl terminus of the second basic amino acid at the RR site in the substrate. The Dibasic Endoprotease Activity Assay Kit uses the synthetic substrate Boc-QRR-pNA (Cat#20820ES) containing a specific peptide-pNA design. This substrate has no absorbance at 405 nm. When Kex2 protease cleaves at the RR site, two fragments are produced: Boc-Gln-Arg-Arg and pNA. pNA is deprotonated in alkaline buffer to form p-nitrophenolate (bright yellow, maximum absorbance at 405 nm). Within a certain concentration range, the rate of change in absorbance at 405 nm is positively correlated with enzyme activity, allowing direct quantification of Kex2 catalytic activity.

The Dibasic Endoprotease Activity Assay Kit is a colorimetric-based dedicated detection tool for Kex2 activity, which can be used for rapid and accurate activity quantification in enzyme preparation R&D, production quality control, and applied research.

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Figure

Figure 1. Kex2 activity assay kinetic curve.