Identification of two types binding modes using reverse or diagonal electrophoretic zymography

Binding mode was demonstrated by Brown and Lantz. They demonstrated that enzymes bind to a fixed substrate in SDS-polyacrylamide gels and they do not migrate into the separating gel and hydrolyze the substrate. We found two types of binding modes. One was formed because of binding between the enzyme and substrate, and the other was due to a high pI value (8.8) of the enzyme. These two types were identified using reverse zymography and diagonal electrophoretic zymography. We identified two different samples showing the binding mode; the fruiting body of the oyster mushroom and culture supernatant from Bacillus sp. strain.