Enzymatic Properties of Two Catalytic Modules of Clostridium stercorarium Pectate Lyase Pel9A

Abstract

Clostridium stercorarium F-9 pectate lyase Pel9A is a modular enzyme composed of two hypothetical family-9 catalytic modules of the polysaccharide lyases, CM9-1 and CM9-2, in order from the N terminus. In this study, we constructed and characterized CM9-1 and CM9-2 polypeptides as rCM9-1 and rCM9-2 respectively. Both of them, like the full-length Pel9A, required the Ca²⁺ ion for their enzyme activities and showed high activity toward polygalacturonic acid but lower activity toward pectin. The specific activity of rCM9-2 was three times higher than that of rCM9-1 and rCM9-2 by itself efficiently catalyzed the depolymerization reaction of polygalacturonic acid into monosaccharide as the major product. It was found that rCM9-1 and rCM9-2 adsorbed to polygalacturonic acid and pectin on native affinity PAGE analysis, suggesting that they contain an independent carbohydrate-binding module separable from a catalytic module or consist of a catalytic module with a binding affinity for pectic substrates.