8WG0

Crystal structure of GH97 glucodextranase from Flavobacterium johnsoniae in complex with glucose

Summary for 8WG0

• Entry DOI: 10.2210/pdb8wg0/pdb
• Descriptor: Candidate alpha-glucosidase Glycoside hydrolase family 97, CALCIUM ION, alpha-D-glucopyranose, ... (5 entities in total)
• Functional Keywords: glycoside hydrolase, gh97, dextran, tim-barrel, isomaltose, nigerose, hydrolase
• Biological source: Flavobacterium johnsoniae (strain ATCC 17061 / DSM 2064 / JCM 8514 / NBRC 14942 / NCIMB 11054 / UW101)
• Total number of polymer chains: 4
• Total formula weight: 323437.27

Authors

Kurata, R.,Nakamura, S.,Miyazaki, T. (deposition date: 2023-09-20, release date: 2024-05-08, Last modification date: 2024-07-31)

Primary citation

Nakamura, S.,Kurata, R.,Miyazaki, T.
Structural insights into alpha-(1→6)-linkage preference of GH97 glucodextranase from Flavobacterium johnsoniae.
Febs J., 291:3267-3282, 2024

PubMed Abstract: Glycoside hydrolase family 97 (GH97) comprises enzymes like anomer-inverting α-glucoside hydrolases (i.e., glucoamylase) and anomer-retaining α-galactosidases. In a soil bacterium, Flavobacterium johnsoniae, we previously identified a GH97 enzyme (FjGH97A) within the branched dextran utilization locus. It functions as an α-glucoside hydrolase, targeting α-(1→6)-glucosidic linkages in dextran and isomaltooligosaccharides (i.e., glucodextranase). FjGH97A exhibits a preference for α-(1→6)-glucoside linkages over α-(1→4)-linkages, while Bacteroides thetaiotaomicron glucoamylase SusB (with 69% sequence identity), which is involved in the starch utilization system, exhibits the highest specificity for α-(1→4)-glucosidic linkages. Here, we examined the crystal structures of FjGH97A in complexes with glucose, panose, or isomaltotriose, and analyzed the substrate preferences of its mutants to identify the amino acid residues that determine the substrate specificity for α-(1→4)- and α-(1→6)-glucosidic linkages. The overall structure of FjGH97A resembles other GH97 enzymes, with conserved catalytic residues similar to anomer-inverting GH97 enzymes. A comparison of active sites between FjGH97A and SusB revealed differences in amino acid residues at subsites +1 and +2 (specifically Ala195 and Ile378 in FjGH97A). Among the three mutants (A195S, I378F, and A195S-I378F), A195S and A195S-I378F exhibited increased activity toward α-(1→4)-glucoside bonds compared to α-(1→6)-glucoside bonds. This suggests that Ala195, located on the Gly184-Thr203 loop (named loop-N) conserved within the GH97 subgroup, including FjGH97A and SusB, holds significance in determining linkage specificity. The conservation of alanine in the active site of the GH97 enzymes, within the same gene cluster as the putative dextranase, indicates its crucial role in determining the specificity for α-(1→6)-glucoside linkage.

PubMed: 38661728
DOI: 10.1111/febs.17139

Experimental method

X-RAY DIFFRACTION (1.95 Å)