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Structural and thermodynamic dissection of specific mannan recognition by a carbohydrate binding module, TmCBM27
Alisdair B Boraston1, Timothy J Revett, Catherine M Boraston
1Department of Chemistry, University of York, Heslington, York YO10 5YW, United Kingdom.
Abstract:
The C-terminal 176 amino acids of a Thermotoga maritima mannanase (Man5) constitute a carbohydrate binding module (CBM) that has been classified into CBM family 27. The isolated CBM27 domain, named TmCBM27, binds tightly (K(a)s 10(5)-10(6) M(-1)) to beta-1, 4-mannooligosaccharides, carob galactomannan, and konjac glucomannan, but not to cellulose (insoluble and soluble) or soluble birchwood xylan. The X-ray crystal structures of native TmCBM27, a TmCBM27-mannohexaose complex, and a TmCBM27-6(3),6(4)-alpha-D-galactosyl-mannopentaose complex at 2.0 A, 1.6 A, and 1.35 A, respectively, reveal the basis of TmCBM27's specificity for mannans. In particular, the latter complex, which is the first structure of a CBM in complex with a branched plant cell wall polysaccharide, illustrates how the architecture of the binding site can influence the recognition of naturally substituted polysaccharides.
Insights
The Thermotoga maritima mannanase CBM27 domain specifically binds mannans, not cellulose or xylan. Structural analysis reveals how its binding site architecture recognizes branched mannans.
Area of Science:
- Biochemistry
- Structural Biology
- Carbohydrate Chemistry
Background:
- Mannanases are enzymes that degrade mannans, important components of plant cell walls.
- Carbohydrate-binding modules (CBMs) are crucial for enzyme targeting and substrate recognition.
- CBM family 27 (CBM27) is known to interact with mannans, but its structural basis for specificity is not fully understood.
Purpose of the Study:
- To characterize the binding properties and specificity of the CBM27 domain from Thermotoga maritima mannanase (TmCBM27).
- To elucidate the structural basis for TmCBM27's recognition of mannooligosaccharides and branched mannans through X-ray crystallography.
Main Methods:
- Isolation and purification of the TmCBM27 domain.
- Binding assays to determine affinity for various polysaccharides (mannans, cellulose, xylan).
- X-ray crystallography to determine the structures of native TmCBM27 and its complexes with ligands.
Main Results:
- TmCBM27 exhibits high affinity for beta-1,4-mannooligosaccharides, carob galactomannan, and konjac glucomannan.
- TmCBM27 does not bind to cellulose or xylan, indicating high specificity.
- Crystal structures reveal the binding site architecture and how it accommodates mannooligosaccharides and branched galactomannans.
Conclusions:
- TmCBM27 is a specific mannan-binding CBM.
- The study provides the first structural insights into CBM recognition of branched plant cell wall polysaccharides.
- The binding site architecture dictates the specificity for mannans, including substituted variants.
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