A β-Mannanase with a Lysozyme-like Fold and a Novel Molecular Catalytic Mechanism

Jin, Yi; Petricevic, Marija; John, Alan; Raich, Lluís; Jenkins, Huw T.; Souza, Leticia Portela De; Cuskin, Fiona; Gilbert, Harry J.; Rovira, Carme; Goddard‐Borger, Ethan D.; Williams, Spencer J.; Davies, G.J.

doi:10.1021/acscentsci.6b00232

Cited by 40 publications

(24 citation statements)

References 37 publications

(69 reference statements)

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“…S6A). On the other hand, structures of GH families 5, 26, and 134 ␤-1,4-mannanases display an "open" groove, which has enough space to accommodate the substrates with galactose side chains (13,23,24). Therefore, it can be deduced that enzyme activity of GH family 113 members is limited by the ␣-1,6-galactose residues because of that structural difference.…”

Section: Discussionmentioning

confidence: 99%

Structural insights into the catalytic mechanism of a novel glycoside hydrolase family 113 β-1,4-mannanase from Amphibacillus xylanus

You

Qin

Yan

et al. 2018

Journal of Biological Chemistry

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β-1,4-Mannanase degrades β-1,4-mannan polymers into manno-oligosaccharides with a low degree of polymerization. To date, only one glycoside hydrolase (GH) family 113 β-1,4-mannanase, from (ManA), has been structurally characterized, and no complex structure of enzyme-manno-oligosaccharides from this family has been reported. Here, crystal structures of a GH family 113 β-1,4-mannanase from (Man113A) and its complexes with mannobiose, mannotriose, mannopentaose, and mannahexaose were solved. Man113A had higher affinity for -1 and +1 mannoses, which explains why the enzyme can hydrolyze mannobiose. At least six subsites (-4 to +2) exist in the groove, but mannose units preferentially occupied subsites -4 to -1 because of steric hindrance formed by Lys-238 and Trp-239. Based on the structural information and bioinformatics, rational design was implemented to enhance hydrolysis activity. Enzyme activity ofMan113A mutants V139C, N237W, K238A, and W239Y was improved by 93.7, 63.4, 112.9, and 36.4%, respectively, compared with the WT. In addition, previously unreported surface-binding sites were observed. Site-directed mutagenesis studies and kinetic data indicated that key residues near the surface sites play important roles in substrate binding and recognition. These first GH family 113 β-1,4-mannanase-manno-oligosaccharide complex structures may be useful in further studying the catalytic mechanism of GH family 113 members, and provide novel insight into protein engineering of GHs to improve their hydrolysis activity.

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Section: Discussionmentioning

confidence: 99%

Structural insights into the catalytic mechanism of a novel glycoside hydrolase family 113 β-1,4-mannanase from Amphibacillus xylanus

You

Qin

Yan

et al. 2018

Journal of Biological Chemistry

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“…It correlates with the mannose configuration bound to the Ϫ2 subsite of the ␤-mannanase from Cellvibrio japonicus (PDB code 2VX5) (32) and with the mannosyl residues from the mannotriose bound to Streptomyces sp. ␤-mannanase (PDB code 5JU9) (31). In GHs, the substrate saccharide unit that binds to the negative subsite generally adopts a boat-or skew-boat-type conformation before the cleavage (30), such as in retaining ␤-mannosidases/ mannanases that often use the 1 S 5 to B 2,5 to 0 S 2 (33) or the 1 C 4 to 3 H 4 to 3 S 1 (31) catalytic pathways.…”

Section: Gh2 Exo-␤-mannanasementioning

confidence: 99%

Structural basis of exo-β-mannanase activity in the GH2 family

Domingues

Souza

Vieira

et al. 2018

Journal of Biological Chemistry

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The classical microbial strategy for depolymerization of β-mannan polysaccharides involves the synergistic action of at least two enzymes, endo-1,4-β-mannanases and β-mannosidases. In this work, we describe the first exo-β-mannanase from the GH2 family, isolated from pv. (XacMan2A), which can efficiently hydrolyze both manno-oligosaccharides and β-mannan into mannose. It represents a valuable process simplification in the microbial carbon uptake that could be of potential industrial interest. Biochemical assays revealed a progressive increase in the hydrolysis rates from mannobiose to mannohexaose, which distinguishes XacMan2A from the known GH2 β-mannosidases. Crystallographic analysis indicates that the active-site topology of XacMan2A underwent profound structural changes at the positive-subsite region, by the removal of the physical barrier canonically observed in GH2 β-mannosidases, generating a more open and accessible active site with additional productive positive subsites. Besides that, XacMan2A contains two residue substitutions in relation to typical GH2 β-mannosidases, Gly and Gly, which alter the active site volume and are essential to its mode of action. Interestingly, the only other mechanistically characterized mannose-releasing exo-β-mannanase so far is from the GH5 family, and its mode of action was attributed to the emergence of a blocking loop at the negative-subsite region of a cleft-like active site, whereas in XacMan2A, the same activity can be explained by the removal of steric barriers at the positive-subsite region in an originally pocket-like active site. Therefore, the GH2 exo-β-mannanase represents a distinct molecular route to this rare activity, expanding our knowledge about functional convergence mechanisms in carbohydrate-active enzymes.

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“…158,159 This is supported by a broader understanding of the tertiary structures of enzyme targets and the further uncovering of novel mechanisms in this family. 98,160,161 Epoxides such as the natural product cyclophellitol and sulfonium ions like ones found naturally within the salicinol family were both discovered as synthetic classes of glycosidase inhibitor before they were identified in nature. 84,146 The irreversible nature of epoxide inhibitors has recently allowed detection of proteins fused to a bglucosidase in a cellular context.…”

Section: Resultsmentioning

confidence: 99%

Back to (non-)Basics: An Update on Neutral and Charge-Balanced Glycosidase Inhibitors

Simone

Mares

Eveleens

et al. 2018

MRMC

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Glycosidases have important anti-cancer, anti-viral and anti-diabetic properties. This review covers the literature in the past 15 years since our initial review in this journal on "neutral" glycosidase inhibitors lacking a basic nitrogen found in iminosugars and azasugars or inhibitors that are neutral by virtue of being "charge-balanced" (zwitterionic). These structurally diverse inhibitors include lactones, lactams, epoxides such as cyclophellitol, and sulfonium ion derivatives of the natural product salacinol. Synthetic efforts toward cyclophillitol, salicinol and derivatives are also highlighted. Importantly, certain metals can inhibit glycosidases and care must be taken to remove residual catalysts from synthetic material to be tested against these enzymes.

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A β-Mannanase with a Lysozyme-like Fold and a Novel Molecular Catalytic Mechanism

Cited by 40 publications

References 37 publications

Structural insights into the catalytic mechanism of a novel glycoside hydrolase family 113 β-1,4-mannanase from Amphibacillus xylanus

Structural insights into the catalytic mechanism of a novel glycoside hydrolase family 113 β-1,4-mannanase from Amphibacillus xylanus

Structural basis of exo-β-mannanase activity in the GH2 family

Back to (non-)Basics: An Update on Neutral and Charge-Balanced Glycosidase Inhibitors

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