The effect of crawfish proteases on inactivation and the hydrolytic cleavage of pectic enzymes

Four proteases ( CP-I, -II, -III and -IV) were isolated and purified from h epatopancreas of crawfish. Based upon their activities with known inhibitor s, they resembled trypsin. All were cross-reactive with polyclonal antibodi es raised against CP-II, indicating they shared structural components. Each of these was able to inactivate orange peel pectinesterase, tomato pectine sterase and pectate lyase C at room temperature under nondenaturing conditi ons. Using matrix-assisted-laser desorption time of flight mass spectrometr y (MALDI-TOF MS), an analysis of peptides generated during proteolysis of p ectate lyase C from Erwinia chrysenthemi showed similar peptide patterns fo r the four crawfish proteases, indicating a common specificity for each iso zyme. However, the cleavage patterns were different from those obtained by the action of bovine trypsin on pectate lyase C These studies indicate the four proteases from the hepatopancreas of crawfish are isozymes which may b e used for the inactivation of pectinolytic enzymes.