7exv
GH127 beta-L-arabinofuranosidase HypBA1 covalently complexed with beta-L-arabinofuranoylamide
Structural highlights
FunctionHYBA1_BIFL2 Beta-L-arabinofuranosidase that removes the beta-L-arabinofuranose residue from the non-reducing end of various substrates, including beta-L-arabinofuranosyl-hydroxyproline (Ara-Hyp), Ara-beta-1,2-Ara-beta-Hyp (Ara(2)-Hyp), Ara-beta-1,2-Ara-beta-1,2-Ara-beta-Hyp (Ara(3)-Hyp), and beta-L-arabinofuranosyl-(1->2)-1-O-methyl-beta-L-arabinofuranose. In the presence of 1-alkanols, shows transglycosylation activity, retaining the anomeric configuration of the arabinofuranose residue. Publication Abstract from PubMedbeta-l-Arabinofuranosidase HypBA1 from Bifidobacterium longum belongs to the glycoside hydrolase family 127. At the active site of HypBA1, a cysteine residue (Cys417) coordinates with a Zn2+ atom and functions as the catalytic nucleophile for the anomer-retaining hydrolytic reaction. In this study, the role of Zn2+ ion and cysteine in catalysis as well as the substrate-bound structure were studied based on biochemical and crystallographic approaches. The enzymatic activity of HypBA1 decreased after dialysis in the presence of EDTA and guanidine hydrochloride and was then recovered by the addition of Zn2+. The Michaelis complex structure was determined using a crystal of a mutant at the acid/base catalyst residue (E322Q) soaked in a solution containing the substrate p-nitrophenyl-beta-l-arabinofuranoside. To investigate the covalent thioglycosyl enzyme intermediate structure, synthetic inhibitors of l-arabinofuranosyl haloacetamide derivatives with different anomer configurations were used to target the nucleophilic cysteine. In the crystal structure of HypBA1, beta-configured l-arabinofuranosylamide formed a covalent link with Cys417, whereas alpha-configured l-arabinofuranosylamide was linked to a noncatalytic residue Cys415. Mass spectrometric analysis indicated that Cys415 was also reactive with the probe molecule. With the beta-configured inhibitor, the arabinofuranoside moiety was correctly positioned at the subsite and the active site integrity was retained to successfully mimic the covalent intermediate state. Substrate complex structure, active site labeling and catalytic role of the zinc ion in cysteine glycosidase.,Maruyama S, Sawano K, Amaki S, Suzuki T, Narita S, Kimura K, Arakawa T, Yamada C, Ito Y, Dohmae N, Fujita K, Ishiwata A, Fushinobu S Glycobiology. 2021 Oct 7. pii: 6382528. doi: 10.1093/glycob/cwab103. PMID:34735571[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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