Carbohydrate-binding architecture of the multi-modular α-1,6-glucosyltransferase from Paenibacillus sp. 598K, which produces α-1,6-glucosyl-α-glucosaccharides from starch

Abstract

sp. 598K α-1,6-glucosyltransferase (Ps6TG31A), a member of glycoside hydrolase family 31, catalyzes exo-α-glucohydrolysis and transglucosylation and produces α-1,6-glucosyl-α-glucosaccharides from α-glucan via its disproportionation activity. The crystal structure of Ps6TG31A was determined by an anomalous dispersion method using a terbium derivative. The monomeric Ps6TG31A consisted of one catalytic (β/α)-barrel domain and six small domains, one on the N-terminal and five on the C-terminal side. The structures of the enzyme complexed with maltohexaose, isomaltohexaose, and acarbose demonstrated that the ligands were observed in the catalytic cleft and the sugar-binding sites of four β-domains. The catalytic site was structured by a glucose-binding pocket and an aglycon-binding cleft built by two sidewalls. The bound acarbose was located with its non-reducing end pseudosugar docked in the pocket, and the other moieties along one sidewall serving three subsites for the α-1,4-glucan. The bound isomaltooligosaccharide was found on the opposite sidewall, which provided the space for the acceptor molecule to be positioned for attack of the catalytic intermediate covalent complex during transglucosylation. The N-terminal domain recognized the α-1,4-glucan in a surface-binding mode. Two of the five C-terminal domains belong to the carbohydrate-binding modules family 35 and one to family 61. The sugar complex structures indicated that the first family 35 module preferred α-1,6-glucan, whereas the second family 35 module and family 61 module preferred α-1,4-glucan. Ps6TG31A appears to have enhanced transglucosylation activity facilitated by its carbohydrate-binding modules and substrate-binding cleft that positions the substrate and acceptor sugar for the transglucosylation.

Structural insights into polysaccharide recognition by Flavobacterium johnsoniae dextranase, a member of glycoside hydrolase family 31

“…The crystal structure of GH31 6‐α‐glucosyltransferase from Paenibacillus sp. 598K (Ps6TG31A) has also been reported . Ps6TG31A shows transglucosylation activity against isomaltooligosaccharides and shares some resemblance to FjDex31A, as both enzymes catalyze transglucosylation to form the α‐1,6‐linkage.…”

“…Structural analysis of Ps6TG31A in complex with isomaltohexaose as well as acarbose, an inhibitor of α‐glucosidases, has revealed that Ps6TG31A has multiple substrate‐binding sites . Acarbose binds to subsites −1, +1, +2, and +3 while the β‐isomaltotetraose unit binds to subsites +1a, +2a, +3a, and +4a.…”

“…Many GH31 enzymes are identified as α‐glucosidases that liberate α‐ d ‐glucose from maltooligosaccharides . GH31 also contains various other types of members, including α‐1,3‐glucosidase , α‐xylosidase , α‐galactosidase , maltase‐glucoamylase , sucrase‐isomaltase , α‐glucan lyase , and some glucosyltransferases . Although divergent members are classified into GH31, most GH31 enzymes act on disaccharides and relatively small oligosaccharides in an exo‐acting manner.…”

Crystal structure of GH43 exo-β-1,3-galactanase from the basidiomycetePhanerochaete chrysosporiumprovides insights into the mechanism of bypassing side chains

“…1). The N-terminal catalytic domain consists of a five-bladed b-propeller (Gln21-Gly325), as in other GH clan-F enzymes, and the C-terminal domain (PcCBM35) takes a β-jellyroll fold (Thr326-Tyr448) structure, as in previously reported CBM35s (16)(17)(18)(19)(20)(21)(22)(23)(24)(25).…”

Structural insights into substrate recognition and catalysis by glycoside hydrolase family 87 α‐1,3‐glucanase from Paenibacillus glycanilyticus FH11

“…Although there was very little similarity between the sequence of this module and that of other characterized proteins, proteins structurally similar to this structure were found in the PDB using the DALI server including a number of carbohydrate‐binding module family 35 (CBM35) , including CBM35 of Paenibacillus sp. 598K α‐1,6‐glucosyltransferase (PsCBM35‐1; RMSD = 1.9 Å for the 112 Cα atoms, a.a. sequence identity = 18.8%; PDB: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5X7O) , CBM35 of B. circulans T‐3040 cycloisomaltooligosaccharide glucanotransferase (BcCBM35‐1; RMSD = 2.1 Å for the 117 Cα atoms, a.a. sequence identity = 21.4%; PDB: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=3WNO) , and CBM35 of Clostridium thermocellum (CtCBM35; RMSD = 1.9 Å for the 113 Cα atoms, a.a. sequence identity = 18.6%; PDB: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2WZ8) . CBM35s are accompanied by catalytic domains, including those for xylanase, glucanase, glucosaminidase, mannanase, and galactanase activities, and function to bind their substrate saccharides to assist the catalytic domains.…”