Transition state analysis of Vibrio cholerae sialidase-catalyzed hydrolyses of natural substrate analogues

Download
  1. Get@NRC: Transition state analysis of Vibrio cholerae sialidase-catalyzed hydrolyses of natural substrate analogues (Opens in a new window)
DOIResolve DOI: http://doi.org/10.1021/ja208564y
AuthorSearch for: ; Search for: ; Search for: ; Search for: ; Search for: ; Search for:
TypeArticle
Journal titleJournal of the American Chemical Society
ISSN0002-7863
1520-5126
Volume134
Issue8
Pages37483757; # of pages: 10
AbstractA series of isotopically labeled natural substrate analogues (phenyl 5-N-acetyl-α-d-neuraminyl-(2→3)-β-d-galactopyranosyl-(1→4)-1-thio-β-d-glucopyranoside; Neu5Acα2,3LacβSPh, and the corresponding 2→6 isomer) were prepared chemoenzymatically in order to characterize, by use of multiple kinetic isotope effect (KIE) measurements, the glycosylation transition states for Vibrio cholerae sialidase-catalyzed hydrolysis reactions. The derived KIEs for Neu5Acα2,3LacβSPh for the ring oxygen (18V/K), leaving group oxygen (°⁶V/K), C3-S deuterium (DV/KS) and C3-R deuterium (DV/KR) are 1.029 ± 0.002, 0.983 ± 0.001, 1.034 ± 0.002, and 1.043 ± 0.002, respectively. In addition, the KIEs for Neu5Acα2,6βSPh for C3-S deuterium (DV/KS) and C3-R deuterium (DV/KR) are 1.021 ± 0.001 and 1.049 ± 0.001, respectively. The glycosylation transition state structures for both Neu5Acα2,3LacβSPh and Neu5Acα2,6LacβSPh were modeled computationally using the experimental KIE values as goodness of fit criteria. Both transition states are late with largely cleaved glycosidic bonds coupled to pyranosyl ring flattening (4H5 half-chair conformation) with little or no nucleophilic involvement of the enzymatic tyrosine residue. Notably, the transition state for the catalyzed hydrolysis of Neu5Acα2,6βSPh appears to incorporate a lesser degree of general-acid catalysis, relative to the 2,3-isomer.
Publication date
LanguageEnglish
AffiliationNRC Institute for Biological Sciences; National Research Council Canada
Peer reviewedYes
NPARC number21269000
Export citationExport as RIS
Report a correctionReport a correction
Record identifier7c867330-319e-43c0-859e-77d22ac05b0a
Record created2013-11-29
Record modified2016-05-09
Bookmark and share
  • Share this page with Facebook (Opens in a new window)
  • Share this page with Twitter (Opens in a new window)
  • Share this page with Google+ (Opens in a new window)
  • Share this page with Delicious (Opens in a new window)