Dataset Information

Modulation of activity by Arg407: structure of a fungal alpha-1,2-mannosidase in complex with a substrate analogue.

ABSTRACT: Class I alpha-mannosidases (glycoside hydrolase family GH47) play key roles in the maturation of N-glycans and the ER-associated degradation of unfolded glycoproteins. The 1.95 A resolution structure of a fungal alpha-1,2-mannosidase in complex with the substrate analogue methyl-alpha-D-lyxopyranosyl-(1',2)-alpha-D-mannopyranoside (LM) shows the intact disaccharide spanning the -1/+1 subsites, with the D-lyxoside ring in the -1 subsite in the 1C4 chair conformation, and provides insight into the mechanism of catalysis. The absence of the C5' hydroxymethyl group on the D-lyxoside moiety results in the side chain of Arg407 adopting two alternative conformations: the minor one interacting with Asp375 and the major one interacting with both the D-lyxoside and the catalytic base Glu409, thus disrupting its function. Chemical modification of Asp375 has previously been shown to inactivate the enzyme. Taken together, the data suggest that Arg407, which belongs to the conserved sequence motif RPExxE, may act to modulate the activity of the enzyme. The proposed mechanism for modulating the activity is potentially a general mechanism for this superfamily.

SUBMITTER: Lobsanov YD

PROVIDER: S-EPMC2430470 | biostudies-literature | 2008 Mar

REPOSITORIES: biostudies-literature

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Modulation of activity by Arg407: structure of a fungal alpha-1,2-mannosidase in complex with a substrate analogue.

Lobsanov Yuri D YD Yoshida Takashi T Desmet Tom T Nerinckx Wim W Yip Patrick P Claeyssens Marc M Herscovics Annette A Howell P Lynne PL

Acta crystallographica. Section D, Biological crystallography 20080220 Pt 3

Class I alpha-mannosidases (glycoside hydrolase family GH47) play key roles in the maturation of N-glycans and the ER-associated degradation of unfolded glycoproteins. The 1.95 A resolution structure of a fungal alpha-1,2-mannosidase in complex with the substrate analogue methyl-alpha-D-lyxopyranosyl-(1',2)-alpha-D-mannopyranoside (LM) shows the intact disaccharide spanning the -1/+1 subsites, with the D-lyxoside ring in the -1 subsite in the 1C4 chair conformation, and provides insight into the ...[more]

PMID: 18323617

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Project description:The enzymatic hydrolysis of alpha-mannosides is catalyzed by glycoside hydrolases (GH), termed alpha-mannosidases. These enzymes are found in different GH sequence-based families. Considerable research has probed the role of higher eukaryotic "GH38" alpha-mannosides that play a key role in the modification and diversification of hybrid N-glycans; processes with strong cellular links to cancer and autoimmune disease. The most extensively studied of these enzymes is the Drosophila GH38 alpha-mannosidase II, which has been shown to be a retaining alpha-mannosidase that targets both alpha-1,3 and alpha-1,6 mannosyl linkages, an activity that enables the enzyme to process GlcNAc(Man)(5)(GlcNAc)(2) hybrid N-glycans to GlcNAc(Man)(3)(GlcNAc)(2). Far less well understood is the observation that many bacterial species, predominantly but not exclusively pathogens and symbionts, also possess putative GH38 alpha-mannosidases whose activity and specificity is unknown.Here we show that the Streptococcus pyogenes (M1 GAS SF370) GH38 enzyme (Spy1604; hereafter SpGH38) is an alpha-mannosidase with specificity for alpha-1,3 mannosidic linkages. The 3D X-ray structure of SpGH38, obtained in native form at 1.9 A resolution and in complex with the inhibitor swainsonine (K(i) 18 microM) at 2.6 A, reveals a canonical GH38 five-domain structure in which the catalytic "-1" subsite shows high similarity with the Drosophila enzyme, including the catalytic Zn(2+) ion. In contrast, the "leaving group" subsites of SpGH38 display considerable differences to the higher eukaryotic GH38s; features that contribute to their apparent specificity.Although the in vivo function of this streptococcal GH38 alpha-mannosidase remains unknown, it is shown to be an alpha-mannosidase active on N-glycans. SpGH38 lies on an operon that also contains the GH84 hexosaminidase (Spy1600) and an additional putative glycosidase. The activity of SpGH38, together with its genomic context, strongly hints at a function in the degradation of host N- or possibly O-glycans. The absence of any classical signal peptide further suggests that SpGH38 may be intracellular, perhaps functioning in the subsequent degradation of extracellular host glycans following their initial digestion by secreted glycosidases.

Dataset Information

Modulation of activity by Arg407: structure of a fungal alpha-1,2-mannosidase in complex with a substrate analogue.

Publications

Modulation of activity by Arg407: structure of a fungal alpha-1,2-mannosidase in complex with a substrate analogue.

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