6SXU

GH51 a-l-arabinofuranosidase soaked with cyclic sulfate inhibitor

This is a non-PDB format compatible entry.

Summary for 6SXU

• Entry DOI: 10.2210/pdb6sxu/pdb
• Descriptor: Intracellular exo-alpha-(1->5)-L-arabinofuranosidase, [(1~{S},2~{S},3~{S},4~{S})-2-(hydroxymethyl)-3,4-bis(oxidanyl)cyclopentyl] hydrogen sulfate, DI(HYDROXYETHYL)ETHER, ... (6 entities in total)
• Functional Keywords: covalent complex, arabinofuranosidase, gh51, geobacillus, hydrolase
• Biological source: Geobacillus stearothermophilus
• Total number of polymer chains: 2
• Total formula weight: 115773.08

Authors

McGregor, N.G.S.,Davies, G.J. (deposition date: 2019-09-26, release date: 2020-02-26, Last modification date: 2024-11-06)

Primary citation

McGregor, N.G.S.,Artola, M.,Nin-Hill, A.,Linzel, D.,Haon, M.,Reijngoud, J.,Ram, A.,Rosso, M.N.,van der Marel, G.A.,Codee, J.D.C.,van Wezel, G.P.,Berrin, J.G.,Rovira, C.,Overkleeft, H.S.,Davies, G.J.
Rational Design of Mechanism-Based Inhibitors and Activity-Based Probes for the Identification of Retaining alpha-l-Arabinofuranosidases.
J.Am.Chem.Soc., 142:4648-4662, 2020

PubMed Abstract: Identifying and characterizing the enzymes responsible for an observed activity within a complex eukaryotic catabolic system remains one of the most significant challenges in the study of biomass-degrading systems. The debranching of both complex hemicellulosic and pectinaceous polysaccharides requires the production of α-l-arabinofuranosidases among a wide variety of coexpressed carbohydrate-active enzymes. To selectively detect and identify α-l-arabinofuranosidases produced by fungi grown on complex biomass, potential covalent inhibitors and probes which mimic α-l-arabinofuranosides were sought. The conformational free energy landscapes of free α-l-arabinofuranose and several rationally designed covalent α-l-arabinofuranosidase inhibitors were analyzed. A synthetic route to these inhibitors was subsequently developed based on a key Wittig-Still rearrangement. Through a combination of kinetic measurements, intact mass spectrometry, and structural experiments, the designed inhibitors were shown to efficiently label the catalytic nucleophiles of retaining GH51 and GH54 α-l-arabinofuranosidases. Activity-based probes elaborated from an inhibitor with an aziridine warhead were applied to the identification and characterization of α-l-arabinofuranosidases within the secretome of grown on arabinan. This method was extended to the detection and identification of α-l-arabinofuranosidases produced by eight biomass-degrading basidiomycete fungi grown on complex biomass. The broad applicability of the cyclophellitol-derived activity-based probes and inhibitors presented here make them a valuable new tool in the characterization of complex eukaryotic carbohydrate-degrading systems and in the high-throughput discovery of α-l-arabinofuranosidases.

PubMed: 32053363
DOI: 10.1021/jacs.9b11351

Experimental method

X-RAY DIFFRACTION (1.398 Å)