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499 a.a.
_CL
Key reference
DOI no: 10.1074/jbc.M107154200 J Biol Chem 276:42131-42137 (2001) PubMed id: 11522797
Biochemical and structural assessment of the 1-N-azasugar GalNAc-isofagomine as a potent family 20 beta-N-acetylhexosaminidase inhibitor. B.L.Mark, D.J.Vocadlo, D.Zhao, S.Knapp, S.G.Withers, M.N.James. ABSTRACT
Azasugar inhibitors of the isofagomine class are potent competitive inhibitors of configuration-retaining beta-glycosidases. This potency results from the formation of a strong electrostatic interaction between a protonated endocyclic nitrogen at the "anomeric" center of the inhibitor and the catalytic nucleophile of the enzyme. Although the majority of retaining beta-glycosidases use a mechanism involving a carboxylate residue as a nucleophile, Streptomyces plicatus beta-N-acetylhexos-aminidase (SpHEX) and related family 20 glycosidases lack such a catalytic residue and use instead the carbonyl oxygen of the 2-acetamido group of the substrate as a nucleophile to "attack" the anomeric center. Thus, a strong electrostatic interaction between the inhibitor and enzyme is not expected to occur; nonetheless, the 1-N-azasugar (2R,3R,4S,5R)-2-acetamido-3,4-dihydroxy-5-hydroxymethyl-piperidinium hydrochloride (GalNAc-isofagomine.HCl), which was synthesized and assayed for its ability to inhibit SpHEX, was found to be a potent competitive inhibitor of the enzyme (K(i) = 2.7 microm). A crystallographic complex of GalNAc-isofagomine bound to SpHEX was solved and refined to 1.75 A and revealed that the lack of a strong electrostatic interaction between the "anomeric" center of GalNAc-isofagomine and SpHEX is compensated for by a novel 2.8-A hydrogen bond formed between the equatorial proton of the endocyclic nitrogen of the azasugar ring and the carboxylate of the general acid-base residue Glu-314 of SpHEX. This interaction appears to contribute to the unexpected potency of GalNAc-isofagomine toward SpHEX.
Selected figure(s)
Figure 3.
Fig. 3. Inhibition of SpHEX-catalyzed hydrolysis of pNPGlcNAc by GalNAc-isofagomine (3, Fig. 2). The concentrations of GalNAc-isofagomine were 0.62 (circle ), 1.23 (
), 3.08 (
), 6.15 (
), 9.23 (
), and 18.5 (
) µM.
Figure 5.
Fig. 5. Schematic of the hydrogen-bonding interactions occurring between SpHEX and GalNAc-isofagomine (3, Fig. 2). Hydrogen bonds are shown as dashed lines, and their respective distances are indicated in Ångstroms. GalNAc-isofagomine is shown in blue, and the SpHEX residue side chains are shown in black and numbered. No attempt has been made to locate the true positions of the amino acid side chains.The above figures are reprinted by permission from the ASBMB: J Biol Chem (2001, 276, 42131-42137) copyright 2001. Figures were selected by an automated process.
Literature references that cite this PDB file's key reference
PubMed id Reference
19275764 H.C.Dorfmueller, V.S.Borodkin, M.Schimpl, and D.M.van Aalten (2009).
GlcNAcstatins are nanomolar inhibitors of human O-GlcNAcase inducing cellular hyper-O-GlcNAcylation.Biochem J, 420, 221-227.
PDB code: 2wb5
18074341 A.D.Hill, and P.J.Reilly (2008).
A Gibbs free energy correlation for automated docking of carbohydrates.J Comput Chem, 29, 1131-1141.
17509134 R.Ettrich, V.Kopecký, K.Hofbauerová, V.Baumruk, P.Novák, P.Pompach, P.Man, O.Plíhal, M.Kutý, N.Kulik, J.Sklenár, H.Ryslavá, V.Kren, and K.Bezouska (2007).
Structure of the dimeric N-glycosylated form of fungal beta-N-acetylhexosaminidase revealed by computer modeling, vibrational spectroscopy, and biochemical studies.BMC Struct Biol, 7, 32.
16762038 C.Mayer, D.J.Vocadlo, M.Mah, K.Rupitz, D.Stoll, R.A.Warren, and S.G.Withers (2006).
Characterization of a beta-N-acetylhexosaminidase and a beta-N-acetylglucosaminidase/beta-glucosidase from Cellulomonas fimi.FEBS J, 273, 2929-2941.
16541109 F.V.Rao, H.C.Dorfmueller, F.Villa, M.Allwood, I.M.Eggleston, and D.M.van Aalten (2006).
Structural insights into the mechanism and inhibition of eukaryotic O-GlcNAc hydrolysis.EMBO J, 25, 1569-1578.
PDB codes: 2cbi 2cbj The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.
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