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Unprecedented glycosidase activity at a lectin carbohydrate-binding site exemplified by the cyanobacterial lectin MVL.

ABSTRACT: Carbohydrate binding proteins, or lectins, are engendered with the ability to bind specific carbohydrate structures, thereby mediating cell-cell and cell-pathogen interactions. Lectins are distinct from carbohydrate modifying enzymes and antibodies, respectively, as they do not carry out glycosidase or glycosyl transferase reactions, and they are of nonimmune origin. Cyanobacterial and algal lectins have become prominent in recent years due to their unique biophysical traits, such as exhibiting novel protein folds and unusually high carbohydrate affinity, and ability to potently inhibit HIV-1 entry through high affinity carbohydrate-mediated interactions with the HIV envelope glycoprotein gp120. The antiviral cyanobacterial lectin Microcystis viridis lectin (MVL), which contains two high affinity oligomannose binding sites, is one such example. Here we used glycan microarray profiling, NMR spectroscopy, and mutagenesis to show that one of the two oligomannose binding sites of MVL can catalyze the cleavage of chitin fragments (such as chitotriose) to GlcNAc, to determine the mode of MVL binding to and cleavage of chitotriose, to identify Asp75 as the primary catalytic residue involved in this cleavage, and to solve the solution structure of an inactive mutant of MVL in complex with this unexpected substrate. These studies represent the first demonstration of dual catalytic activity and carbohydrate recognition for discrete oligosaccharides at the same carbohydrate-binding site in a lectin. Sequence comparisons between the N- and C-domains of MVL, together with the sequences of new MVL homologues identified through bioinformatics, provide insight into the evolving roles of carbohydrate recognition.

SUBMITTER: Shahzad-ul-Hussan S

PROVIDER: S-EPMC2792741 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

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Unprecedented glycosidase activity at a lectin carbohydrate-binding site exemplified by the cyanobacterial lectin MVL.

Shahzad-ul-Hussan Syed S Cai Mengli M Bewley Carole A CA

Journal of the American Chemical Society 20091101 45

Carbohydrate binding proteins, or lectins, are engendered with the ability to bind specific carbohydrate structures, thereby mediating cell-cell and cell-pathogen interactions. Lectins are distinct from carbohydrate modifying enzymes and antibodies, respectively, as they do not carry out glycosidase or glycosyl transferase reactions, and they are of nonimmune origin. Cyanobacterial and algal lectins have become prominent in recent years due to their unique biophysical traits, such as exhibiting ...[more]

PMID: 19856962

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Project description:BackgroundLectins have come a long way from being identified as proteins that agglutinate cells to promising therapeutic agents in modern medicine. Through their specific binding property, they have proven to be anti-cancer, anti-insect, anti-viral agents without affecting the non-target cells. The Arisaema tortuosum lectin (ATL) is a known anti-insect and anti-cancer candidate, also has interesting physical properties. In the present work, its carbohydrate binding behavior is investigated in detail, along with its anti-proliferative property.ResultsThe microcalorimetry of ATL with a complex glycoprotein asialofetuin demonstrated trivalency contributed by multiple binding sites and enthalpically driven spontaneous association. The complex sugar specificity of ATL towards multiple sugars was also demonstrated in glycan array analysis in which the trimannosyl pentasaccharide core N-glycan [Manα1-6(Manα1-3)Manβ1-4GlcNAcβ1-4GlcNAcβ] was the highest binding motif. The high binding glycans for ATL were high mannans, complex N-glycans, core fucosylated N-glycans and glycans with terminal lactosamine units attached to pentasaccharide core. ATL induced cell death in IMR-32 cells was observed as time dependent loss in cell number, formation of apoptotic bodies and DNA damage. As a first report of molecular cloning of ATL, the in silico analysis of its cDNA revealed ATL to be a β-sheet rich heterotetramer. A homology model of ATL showed beta prism architecture in each monomer with 85% residues in favoured region of Ramachandran plot.ConclusionsDetailed exploration of carbohydrate binding behavior indicated ATL specificity towards complex glycans, while no binding to simple sugars, including mannose. Sequence analysis of ATL cDNA revealed that during the tandem evolutionary events, domain duplication and mutations lead to the loss of mannose specificity, acquiring of new sugar specificity towards complex sugars. It also resulted in the formation of a two-domain single chain polypeptide with both domains having different binding sites due to mutations within the consensus carbohydrate recognition sites [QXDXNXVXY]. This unique sugar specificity can account for its significant biological properties. Overall finding of present work signifies anti-cancer, anti-insect and anti-viral potential of ATL making it an interesting molecule for future research and/or theragnostic applications.

Dataset Information

Unprecedented glycosidase activity at a lectin carbohydrate-binding site exemplified by the cyanobacterial lectin MVL.

Publications

Unprecedented glycosidase activity at a lectin carbohydrate-binding site exemplified by the cyanobacterial lectin MVL.

Similar Datasets

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