Harnessing glycosylation to improve cellulase activity

Beckham, Gregg T.; Dai, Ziyu; Matthews, James F.; Momany, Michelle; Payne, Christina M.; Adney, William S.; Baker, Scott E.; Himmel, Michael E.

doi:10.1016/j.copbio.2011.11.030

Cited by 114 publications

(112 citation statements)

References 50 publications

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“…Subsequently, we synthesized two additional series of CBMs that had specific glycans at more than one of three glycosylation sites (11)(12)(13)(14)(15)(16)(17)(18)(19)(20) (Fig. 2).…”

Section: Resultsmentioning

confidence: 99%

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Specificity of O -glycosylation in enhancing the stability and cellulose binding affinity of Family 1 carbohydrate-binding modules

Chen

Drake

Resch

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

154

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The majority of biological turnover of lignocellulosic biomass in nature is conducted by fungi, which commonly use Family 1 carbohydrate-binding modules (CBMs) for targeting enzymes to cellulose. Family 1 CBMs are glycosylated, but the effects of glycosylation on CBM function remain unknown. Here, the effects of O-mannosylation are examined on the Family 1 CBM from the Trichoderma reesei Family 7 cellobiohydrolase at three glycosylation sites. To enable this work, a procedure to synthesize glycosylated Family 1 CBMs was developed. Subsequently, a library of 20 CBMs was synthesized with mono-, di-, or trisaccharides at each site for comparison of binding affinity, proteolytic stability, and thermostability. The results show that, although CBM mannosylation does not induce major conformational changes, it can increase the thermolysin cleavage resistance up to 50-fold depending on the number of mannose units on the CBM and the attachment site. O-Mannosylation also increases the thermostability of CBM glycoforms up to 16°C, and a mannose disaccharide at Ser3 seems to have the largest themostabilizing effect. Interestingly, the glycoforms with small glycans at each site displayed higher binding affinities for crystalline cellulose, and the glycoform with a single mannose at each of three positions conferred the highest affinity enhancement of 7.4-fold. Overall, by combining chemical glycoprotein synthesis and functional studies, we show that specific glycosylation events confer multiple beneficial properties on Family 1 CBMs. chemical synthesis | cellulase | biofuels | protein engineering

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“…Subsequently, we synthesized two additional series of CBMs that had specific glycans at more than one of three glycosylation sites (11)(12)(13)(14)(15)(16)(17)(18)(19)(20) (Fig. 2).…”

Section: Resultsmentioning

confidence: 99%

“…Glycosylation is an important heterogeneous posttranslational modification (PTM) in fungal enzymes that degrade biomass (16,17). To date, few studies have been conducted to examine glycosylation in secreted fungal enzymes to determine the extent and factors that control it, including growth conditions and extracellular glycan-trimming enzymes.…”

mentioning

confidence: 99%

Specificity of O -glycosylation in enhancing the stability and cellulose binding affinity of Family 1 carbohydrate-binding modules

Chen

Drake

Resch

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

154

View full text Add to dashboard Cite

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“…For the cellodextrin-bound enzymes, we consider two different binding scenarios: a cellobiose bound at the Ϫ2, Ϫ1 subsites and a cellotetraose bound from the ϩ1 to ϩ4 subsites, as shown in the crystal structure, and a single cellohexaose bound in the tunnel from the Ϫ2 to ϩ4 subsites. Additionally, a previously equilibrated cellulose I␤ microfibril (42,43) was used to construct a TfuCel6B-cellulose complex (Fig. 3).…”

Section: Methodsmentioning

confidence: 99%

Loop Motions Important to Product Expulsion in the Thermobifida fusca Glycoside Hydrolase Family 6 Cellobiohydrolase from Structural and Computational Studies

Vuong

et al. 2013

Journal of Biological Chemistry

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Background: Family 6 glycoside hydrolases represent an important, diverse enzyme class in cellulolytic organisms. Results: We solved structures of two Thermobifida fusca Cel6B (TfuCel6B) cellobiohydrolase mutants and examined ligand dynamics and product release with simulation. Conclusion: These results suggest mechanisms for product release in TfuCel6B. Significance: This study further elucidates the mechanism of a unique cellobiohydrolase with an extended, enclosed active site tunnel.

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“…Cellulase consists of three major components, cellobiohydrolase, endoglucanase, and β-glucosidase (Becham et al 2012;Jalak et al 2012). Endoglucanases (EG; Cel5A and Cel7B) and cellobiohydrolases (CBH; Cel6A and Cel7A) are the main cellulase components to depolymerize cellulose chain.…”

Section: Mechanism On the Improvement Effect Of Amphipathic Lignin Dementioning

confidence: 99%

Improvement of Enzymatic Saccharification of Unbleached Cedar Pulp with Amphipathic Lignin Derivatives

Winarni¹,

Oikawa²,

Yamada³

et al. 2013

BioResources

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Synthesized amphipathic lignin derivatives comprised of acidic acid lignin (AL) with poly(ethylene glycol) diglycidyl ether (PEGDE), ethoxy-(2-hydroxy)-propoxy-poly(ethylene glycol) glycidyl ether (EPEG) or dodecyloxy-poly(ethylene glycol) glycidyl ether (DAEO) were added before the enzymatic saccharification of unbleached cedar pulp along with two commercially available cellulases, Meicelase and Genencor GC220. At the same filter paper unit (FPU) dosage, GC220 showed higher sugar yield than Meicelase. The difference was attributed to the composition of processive and non-processive endoglucanase activities per FPU; GC220 had higher such activities than Meicelase. The sugar yield was significantly improved by the addition of the lignin derivatives. In addition, residual activities after the saccharification were maintained at the higher level by their addition than with polyethylene glycol (PEG) 4000. In particular, EPEG-AL yielded the complete recovery of cellulase activity when using 20 FPU/g of substrate. It was found that the lignin derivatives were directly associated with Cel6A, one of cellulase components, whereas PEG 4000 was not. Thus, it is evident that the lignin derivatives are promising agents to improve the enzymatic saccharification of cellulase.

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Harnessing glycosylation to improve cellulase activity

Cited by 114 publications

References 50 publications

Specificity of O -glycosylation in enhancing the stability and cellulose binding affinity of Family 1 carbohydrate-binding modules

Specificity of O -glycosylation in enhancing the stability and cellulose binding affinity of Family 1 carbohydrate-binding modules

Loop Motions Important to Product Expulsion in the Thermobifida fusca Glycoside Hydrolase Family 6 Cellobiohydrolase from Structural and Computational Studies

Improvement of Enzymatic Saccharification of Unbleached Cedar Pulp with Amphipathic Lignin Derivatives

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