Crystal structures of Geobacillus stearothermophilus α-glucuronidase complexed with its substrate and products: Mechanistic implications

Gali Golan, Dalia Shallom, Anna Teplitsky, Galia Zaide, Smadar Shulami, Timor Baasov, Vivian Stojanoff, Andy Thompson, Yuval Shoham*, Gil Shoham

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

61 Scopus citations

Abstract

α-Glucuronidases cleave the α-1,2-glycosidic bond between 4-O-methyl-D-glucuronic acid and short xylooligomers as part of the hemicellulose degradation system. To date, all of the α-glucuronidases are classified as family 67 glycosidases, which catalyze the hydrolysis via the investing mechanism. Here we describe several high resolution crystal structures of the α-glucuronidase (AguA) from Geobacillus stearothermophilus, in complex with its substrate and products. In the complex of AguA with the intact substrate, the 4-O-methyl-D-glucuronic acid sugar ring is distorted into a half-chair conformation, which is closer to the planar conformation required for the oxocarbenium ion-like transition state structure. In the active site, a water molecule is coordinated between two carboxylic acids, in an appropriate position to act as a nucleophile. From the structural data it is likely that two carboxylic acids, Asp364 and Glu 392, activate together the nucleophilic water molecule. The loop carrying the catalytic general acid Glu285 cannot be resolved in some of the structures but could be visualized in its "open" and "closed" (catalytic) conformations in other structures. The protonated state of Glu285 is presumably stabilized by its proximity to the negative charge of the substrate, representing a new variation of substrate-assisted catalysis mechanism.

Original languageAmerican English
Pages (from-to)3014-3024
Number of pages11
JournalJournal of Biological Chemistry
Volume279
Issue number4
DOIs
StatePublished - 23 Jan 2004

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