Crystal Structure of Polygalacturonase from Erwinia carotovora ssp. carotovora

Abstract

The crystal structure of the 40-kDa endo-polygalacturonase from Erwinia carotovora ssp. carotovora was solved by multiple isomorphous replacement and refined at 1.9 Å to a conventional crystallographic R-factor of 0.198 and R free of 0.239. This is the first structure of a polygalacturonase and comprises a 10 turn righthanded parallel ␤-helix domain with two loop regions forming a "tunnel like" substrate-binding cleft. Sequence conservation indicates that the active site of polygalacturonase is between these two loop regions, and comparison of the structure of polygalacturonase with that of rhamnogalacturonase A from Aspergillus aculeatus enables two conserved aspartates, presumed to be catalytic residues, to be identified. An adjacent histidine, in accord with biochemical results, is also seen. A similarity in overall electrostatic properties of the substrate-binding clefts of polygalacturonase and pectate lyase, which bind and cleave the same substrate, polygalacturonic acid, is also revealed.

Structural requirements of endo polygalacturonase for the interaction with PGIP (polygalacturonase-inhibiting protein)

“…33). The ␤-sheet PB2a, starting from the sixth turn of the ␤-helix, is typical of PGs and is absent in other proteins sharing the same overall fold (34). The length of the ␤-strands is generally short (3 to 5 residues); more variable is the length of the turns (T) between ␤-strands.…”

“…1b). This finding is of some interest in view of the polyanionic nature of the substrate and the fact that bacterial pectate lyases and PGs are generally richer in basic amino acids (34,41). It is well known that fungal PGs interact with and are inhibited by plant inhibitors named PGIPs.…”

The crystal structure of polygalacturonase-inhibiting protein (PGIP), a leucine-rich repeat protein involved in plant defense

“…By careful inspection of the Hbonding pattern, B2 may be considered a single sheet formed by the clustering of four shorter sheets that interact together. The presence of the additional sheet B2 places the fold of PGIP2 between the typical LRR structure and the ␤-helical architecture found in pectate lyases and PGs that contain three to four ␤-sheets (30)(31)(32)(33). PGIP2 belongs to the plant-specific LRR subfamily characterized by the consensus sequence Lt͞sGxIP in the region following the conserved ''␤-structure ϩ Asn-ladder'' (23).…”

Hydrolysis of chitin by Pectinex™

“…It may be interesting to speculate at the reasons behind this at the molecular level. Aspartate residue is implicated in the active site of pectinases as a conserved residue [22]. It is noteworthy that a recent paper indicates that carboxyl groups are an essential group for chitinase from A fumigatus [13].…”