Characterization of Pectate Lyase from Alkalitolerant Bacillus sp. YA-14: Its Action Pattern and Active Center

Pectate lyase from alkalitolerant Bacillus sp. YA-14 is an endo-type pectate lyase which acts randomly at the $\alpha$-1, 4-galacturonan linkage, and requires calcium or strontium ions for its activity. The enzyme is active on low methyl esterified pectin, but the activity toward a high methyl esterified substrate is reduced. The apparent Km's of the enzyme toward sodium polygalacturonic acid, polygalacturonic acid, and various pectins such as apple pectin, citrus pectin, and genu pectin are 0.826 mg/ml, 0.685 mg/ml, and 1.14 mg/ml, respectively. The enzyme activity is inhibited by SDS, urea, and sodium azide, but not by various reducing reagents, such as $\beta$-mercaptoethanol, Na-thiosulfate, Na-sulfate, cystein, and L-ascorbic acid. The enzyme is inactivated by N-bromosuccinimide, $I_2, H_2O_2$. PMSF, and iodoacetate. Judging from the results of their inhibition types, we speculate that tryptophan and serine residues are directly involved in enzyme activity, while tyrosine and methionine residues are indirectly involved in its activity.