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Understanding how noncatalytic carbohydrate binding modules can display specificity for xyloglucan.

ABSTRACT: Plant biomass is central to the carbon cycle and to environmentally sustainable industries exemplified by the biofuel sector. Plant cell wall degrading enzymes generally contain noncatalytic carbohydrate binding modules (CBMs) that fulfil a targeting function, which enhances catalysis. CBMs that bind ?-glucan chains often display broad specificity recognizing ?1,4-glucans (cellulose), ?1,3-?1,4-mixed linked glucans and xyloglucan, a ?1,4-glucan decorated with ?1,6-xylose residues, by targeting structures common to the three polysaccharides. Thus, CBMs that recognize xyloglucan target the ?1,4-glucan backbone and only accommodate the xylose decorations. Here we show that two closely related CBMs, CBM65A and CBM65B, derived from EcCel5A, a Eubacterium cellulosolvens endoglucanase, bind to a range of ?-glucans but, uniquely, display significant preference for xyloglucan. The structures of the two CBMs reveal a ?-sandwich fold. The ligand binding site comprises the ?-sheet that forms the concave surface of the proteins. Binding to the backbone chains of ?-glucans is mediated primarily by five aromatic residues that also make hydrophobic interactions with the xylose side chains of xyloglucan, conferring the distinctive specificity of the CBMs for the decorated polysaccharide. Significantly, and in contrast to other CBMs that recognize ?-glucans, CBM65A utilizes different polar residues to bind cellulose and mixed linked glucans. Thus, Gln(106) is central to cellulose recognition, but is not required for binding to mixed linked glucans. This report reveals the mechanism by which ?-glucan-specific CBMs can distinguish between linear and mixed linked glucans, and show how these CBMs can exploit an extensive hydrophobic platform to target the side chains of decorated ?-glucans.

SUBMITTER: Luis AS

PROVIDER: S-EPMC3576085 | biostudies-literature | 2013 Feb

REPOSITORIES: biostudies-literature

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Understanding how noncatalytic carbohydrate binding modules can display specificity for xyloglucan.

Luís Ana S AS Venditto Immacolata I Temple Max J MJ Rogowski Artur A Baslé Arnaud A Xue Jie J Knox J Paul JP Prates José A M JA Ferreira Luís M A LM Fontes Carlos M G A CM Najmudin Shabir S Gilbert Harry J HJ

The Journal of biological chemistry 20121210 7

Plant biomass is central to the carbon cycle and to environmentally sustainable industries exemplified by the biofuel sector. Plant cell wall degrading enzymes generally contain noncatalytic carbohydrate binding modules (CBMs) that fulfil a targeting function, which enhances catalysis. CBMs that bind β-glucan chains often display broad specificity recognizing β1,4-glucans (cellulose), β1,3-β1,4-mixed linked glucans and xyloglucan, a β1,4-glucan decorated with α1,6-xylose residues, by targeting s ...[more]

PMID: 23229556

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Project description:Background:Effective enzymatic degradation of crystalline polysaccharides requires a synergistic cocktail of hydrolytic enzymes tailored to the wide-ranging degree of substrate crystallinity. To accomplish this type of targeted carbohydrate recognition, nature produces multi-modular enzymes, having at least one catalytic domain appended to one or more carbohydrate binding modules (CBMs). The Type B CBM categorization encompasses several families (i.e., protein folds) of CBMs that are generally thought to selectively bind oligomeric polysaccharides; however, a subset of cellulose-specific CBM families (17 and 28) appear to bind non-crystalline cellulose more tightly than oligomers and in a manner that discriminates between surface topology. Results:To provide insight into this unexplained phenomenon, we investigated the molecular-level origins of oligomeric and non-crystalline carbohydrate recognition in cellulose-specific Type B CBMs using molecular dynamics (MD) simulation and free energy calculations. Examining two CBMs from three different families (4, 17, and 28), we describe how protein-ligand dynamics contribute to observed variations in binding affinity of oligomers within the same CBM family. Comparisons across the three CBM families identified factors leading to modified functionality prohibiting competitive binding, despite similarity in sequence and specificity. Using free energy perturbation with Hamiltonian replica exchange MD, we also examined the hypothesis that the open topology of the binding grooves in families 17 and 28 necessitates tight binding of an oligomer, while the more confined family 4 binding groove does not require the same degree of tight binding. Finally, we elucidated the mechanisms of non-crystalline carbohydrate recognition by modeling CBMs complexed with a partially decrystallized cellulose substrate. Molecular simulation provided structural and dynamic data for direct comparison to oligomeric modes of carbohydrate recognition, and umbrella sampling MD was used to determine ligand binding free energy. Comparing both protein-carbohydrate interactions and ligand binding free energies, which were in good agreement with experimental values, we confirmed the hypothesis that family 17 and 28 CBMs bind non-crystalline cellulose and oligomers with different affinities (i.e., high and low). Conclusions:Our study provides an unprecedented level of insight into the complex solid and soluble carbohydrate substrate recognition mechanisms of Type B CBMs, the findings of which hold considerable promise for enhancing lignocellulosic biomass conversion technology and development of plant cell wall probes.

Dataset Information

Understanding how noncatalytic carbohydrate binding modules can display specificity for xyloglucan.

Publications

Understanding how noncatalytic carbohydrate binding modules can display specificity for xyloglucan.

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