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dc.contributor.authorHuang, W.
dc.contributor.authorBoju, L.
dc.contributor.authorTkalec, L.
dc.contributor.authorSu, H.
dc.contributor.authorYang, H.O.
dc.contributor.authorGunay, N.S.
dc.contributor.authorLinhardt, Robert J.
dc.contributor.authorKim, Y.S.
dc.contributor.authorMatte, A.
dc.contributor.authorCygler, M.
dc.date2001
dc.date.accessioned2022-06-27T16:20:27Z
dc.date.available2022-06-27T16:20:27Z
dc.date.issued2001-02-27
dc.identifier.citationActive Site of Chondroitin AC Lyase Revealed by the Structure of Enzyme-Oligosaccharide Complexes and Mutagenesis, W. Huang, L. Boju, L. Tkalec, H. Su, H.O. Yang, N.S. Gunay, R.J. Linhardt, Y.S. Kim, A. Matte, M. Cygler, Biochemistry, 40, 2359-2372, 2001.
dc.identifier.issn62960
dc.identifier.urihttps://doi.org/10.1021/bi0024254
dc.identifier.urihttps://hdl.handle.net/20.500.13015/5861
dc.descriptionBiochemistry, 40, 2359-2372
dc.descriptionNote : if this item contains full text it may be a preprint, author manuscript, or a Gold OA copy that permits redistribution with a license such as CC BY. The final version is available through the publisher’s platform.
dc.description.abstractThe crystal structures of Flavobacterium heparinium chondroitin AC lyase (chondroitinase AC; EC 4.2.2.5) bound to dermatan sulfate hexasaccharide (DShexa), tetrasaccharide (DStetra), and hyaluronic acid tetrasaccharide (HAtetra) have been refined at 2.0, 2.0, and 2.1 Å resolution, respectively. The structure of the Tyr234Phe mutant of AC lyase bound to a chondroitin sulfate tetrasaccharide (CStetra) has also been determined to 2.3 Å resolution. For each of these complexes, four (DShexa and CStetra) or two (DStetra and HAtetra) ordered sugars are visible in electron density maps. The lyase AC DShexa and CStetra complexes reveal binding at four subsites, −2, −1, +1, and +2, within a narrow and shallow protein channel. We suggest that subsites −2 and −1 together represent the substrate recognition area, +1 is the catalytic subsite and +1 and +2 together represent the product release area. The putative catalytic site is located between the substrate recognition area and the product release area, carrying out catalysis at the +1 subsite. Four residues near the catalytic site, His225, Tyr234, Arg288, and Glu371 together form a catalytic tetrad. The mutations His225Ala, Tyr234Phe, Arg288Ala, and Arg292Ala, revealed residual activity for only the Arg292Ala mutant. Structural data indicate that Arg292 is primarily involved in recognition of the N-acetyl and sulfate moieties of galactosamine, but does not participate directly in catalysis. Candidates for the general base, removing the proton attached to C-5 of the glucuronic acid at the +1 subsite, are Tyr234, which could be transiently deprotonated during catalysis, or His225. Tyrosine 234 is a candidate to protonate the leaving group. Arginine 288 likely contributes to charge neutralization and stabilization of the enolate anion intermediate during catalysis.
dc.description.sponsorshipNational Heart, Lung, and Blood Institute
dc.description.urihttps://login.libproxy.rpi.edu/login?url=https://doi.org/10.1021/bi0024254
dc.languageen_US
dc.language.isoENG
dc.relation.ispartofThe Linhardt Research Labs Online Collection
dc.relation.ispartofRensselaer Polytechnic Institute, Troy, NY
dc.relation.ispartofBiochemistry
dc.relation.urihttps://harc.rpi.edu/
dc.subjectBiology
dc.subjectChemistry and chemical biology
dc.subjectChemical and biological engineering
dc.subjectBiomedical engineering
dc.titleActive Site of Chondroitin AC Lyase Revealed by the Structure of Enzyme-Oligosaccharide Complexes and Mutagenesis
dc.typeArticle
dcterms.accessRightshttps://login.libproxy.rpi.edu/login?url=https://doi.org/10.1021/bi0024254
dcterms.isPartOfJournal
dcterms.isVersionOfhttps://doi.org/10.1021/bi0024254
dc.rights.holderIn Copyright : this Item is protected by copyright and/or related rights. You are free to use this Item in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s). https://rightsstatements.org/page/InC/1.0/
dc.creator.identifierhttps://orcid.org/0000-0003-2219-5833
dc.relation.departmentThe Linhardt Research Labs.
dc.relation.departmentThe Shirley Ann Jackson, Ph.D. Center for Biotechnology and Interdisciplinary Studies (CBIS)
rpi.description.pages2359-2372
rpi.description.volume40


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