Understanding How Noncatalytic Carbohydrate Binding Modules Can Display Specificity for Xyloglucan

Luís, Ana S.; Venditto, I.; Temple, Max J.; Rogowski, Artur; Baslé, Arnaud; Xue, Jie; Knox, John; Prates, José A. M.; Ferreira, L.M.A.; Fontes, Carlos M. G. A.; Najmudin, Shabir; Gilbert, Harry J.

doi:10.1074/jbc.m112.432781

Cited by 33 publications

(31 citation statements)

References 44 publications

(70 reference statements)

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“…In the present study we are able to extend these findings to show how a branched xyloglucan oligomer, XXXG, is targeted in the binding cleft of X‐2 L110F. The study also shows a way for a CBM to bind xyloglucan, through interactions with both the backbone and decorations of sugar units of xyloglucan, in agreement with the binding features of two other recently described multi‐specific CBMs …”

Section: Discussionsupporting

confidence: 88%

“…Carbohydrate binding modules can be used in a variety of applications and are excellent systems for studying the molecular factors affecting affinity and specificity in protein–carbohydrate interactions. Several studies on CBMs have increased knowledge of the molecular determinants for protein–carbohydrate specificity Our aim is to increase understanding of the determinants for protein–carbohydrate interactions in order to improve engineering approaches of novel carbohydrate binders with pre‐determined specificity. In a past structural investigation, binding toward linear ligands exerted by a multi‐specific CBM, X‐2 L110F, was compared to the mode of binding of its unmodified mono‐selective precursor, X‐2.…”

Section: Discussionmentioning

confidence: 99%

“…As xyloglucan is an abundant polysaccharide and has an increasing number of industrial applications, finding tools that ease analysis and detection of this branched and heterogeneous polysaccharide are of substantial interest . For the study of xyloglucan, several proteins that can target this polysaccharide have been generated or found in natural sources, for example CBM44 and CBM30, CBM62 from Clostridium thermocellum , or CBM65s derived from Eubacterium cellulosolvens Cel5A . The mode in which xyloglucan is recognized by CBMs can roughly be divided into three categories: (1) CBMs that principally recognize the β‐1,4‐glucose backbone and do not display a definitive preference for xyloglucan over unbranched β‐1,4‐glucose polymers; (2) CBMs that mainly target the branching sugar units of xyloglucan such as CBM62, which interacts with β ‐ d ‐galactoses decorated on the xylose sugars of xyloglucan; (3) CBMs, such as X‐2 L110F and CBM65B of Ec Cel5A, which interact with both the β‐1,4‐glucose backbone and the xylose side chains, reflected by a preference for xyloglucan over unbranched β‐1,4‐glucose polymers.…”

Section: Discussionmentioning

confidence: 99%

“…For instance, the affinity of X‐2 L110F for oat‐spelt xylan is about 3.5 times higher than its affinity for tamarind seed xyloglucan . In contrast, the affinity of CBM65B for a variety of xylans is minimal . The inability of CBM65s to recognize xylans is believed to be caused by the lack of C6 atoms in xylose sugars that can interact with the CBM in the same way glucoses on XXXG do and also due to the architecture of the CBM65 binding cleft, which is not optimized for binding xylan, which often adopts a 120° screw axis conformation .…”

Section: Discussionmentioning

confidence: 99%

“…For CBMs, including CBM65s, the position of aromatic side chains in the binding cleft has also been suggested to be of high significance for interaction with carbohydrates. In particular aromatic side chains situated in central positions in the cleft, such as F110 in X‐2 L110F, appear to have a more pronounced role in ligand binding. From this present study and our previous one we conclude that the importance of the central aromatic amino acid F110 in X‐2 L110F depends on the capacity of this side chain to be realigned and bind to CH groups of various ligands by providing a flexible platform for CH‐π interactions, thus affording the CBM the ability to cross‐react with several different carbohydrates, including branched polymers such as xyloglucan.…”

Section: Discussionmentioning

confidence: 99%

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Carbohydrate binding module recognition of xyloglucan defined by polar contacts with branching xyloses and CH‐Π interactions

et al. 2014

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Engineering of novel carbohydrate-binding proteins that can be utilized in various biochemical and biotechnical applications would benefit from a deeper understanding of the biochemical interactions that determine protein-carbohydrate specificity. In an effort to understand further the basis for specificity we present the crystal structure of the multi-specific carbohydrate-binding module (CBM) X-2 L110F bound to a branched oligomer of xyloglucan (XXXG). X-2 L110F is an engineered CBM that can recognize xyloglucan, xylans and β-glucans. The structural observations of the present study compared with previously reported structures of X-2 L110F in complex with linear oligomers, show that the π-surface of a phenylalanine, F110, allows for interactions with hydrogen atoms on both linear (xylopentaose and cellopentaose) and branched ligands (XXXG). Furthermore, X-2 L110F is shown to have a relatively flexible binding cleft, as illustrated in binding to XXXG. This branched ligand requires a set of reorientations of protein side chains Q72, N31, and R142, although these residues have previously been determined as important for binding to xylose oligomers by mediating polar contacts. The loss of these polar contacts is compensated for in binding to XXXG by polar interactions mediated by other protein residues, T74, R115, and Y149, which interact mainly with the branching xyloses of the xyloglucan oligomer. Taken together, the present study illustrates in structural detail how CH-π interactions can influence binding specificity and that flexibility is a key feature for the multi-specificity displayed by X-2 L110F, allowing for the accommodation of branched ligands.

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Section: Discussionsupporting

confidence: 88%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Carbohydrate binding module recognition of xyloglucan defined by polar contacts with branching xyloses and CH‐Π interactions

et al. 2014

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Advances in molecular engineering of carbohydrate-binding modules

et al. 2017

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Carbohydrate-binding modules (CBMs) are non-catalytic domains that are generally appended to carbohydrate-active enzymes. CBMs have a broadly conserved structure that allows recognition of a notable variety of carbohydrates, in both their soluble and insoluble forms, as well as in their alpha and beta conformations and with different types of bonds or substitutions. This versatility suggests a high functional plasticity that is not yet clearly understood, in spite of the important number of studies relating protein structure and function. Several studies have explored the flexibility of these systems by changing or improving their specificity toward substrates of interest. In this review, we examine the molecular strategies used to identify CBMs with novel or improved characteristics. The impact of the spatial arrangement of the functional amino acids of CBMs is discussed in terms of unexpected new functions that are not related to the original biological roles of the enzymes. Proteins 2017; 85:1602-1617. © 2017 Wiley Periodicals, Inc.

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