Feruloyl Esterase Activity of the
            <i>Clostridium thermocellum</i>
            Cellulosome Can Be Attributed to Previously Unknown Domains of XynY and XynZ

Blum, David; Kataeva, Irina; Li, Xinliang; Ljungdahl, Lars G.

doi:10.1128/jb.182.5.1346-1351.2000

Cited by 139 publications

(107 citation statements)

References 42 publications

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“…Xylanase Z from C. thermocellum (XynZt) was selected as the hemicellulase of choice for this work because this very potent enzyme possesses an interacting pair of complementary catalytic modules, i.e. a family-10 xylanase and a feruloyl esterase (19). In the free state, the three enzymes exhibited relatively low levels of enhanced synergy (approximately 30%) in the degradation of straw (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Among the available C. thermocellum xylanase-encoding genes, the xynZ gene is of particular interest. The corresponding protein is in itself a bifunctional enzyme (19). The enzyme contains a family-1 carbohydrate esterase catalytic module, a family-6 CBM, a typical C. thermocellum type I dockerin module, and a family-10 xylanase catalytic module (afmb.cnrsmrs.fr/CAZY/).…”

Section: Selection and Characterization Of A Third Divergentmentioning

confidence: 99%

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Action of Designer Cellulosomes on Homogeneous Versus Complex Substrates

Fierobe

Mingardon

Mechaly

et al. 2005

Journal of Biological Chemistry

217

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(2002) J. Biol. Chem. 277, 49621-49630), we reported the self-assembly of a comprehensive set of defined "bifunctional" chimeric cellulosomes. Each complex contained the following: (i) a chimeric scaffoldin possessing a cellulose-binding module and two cohesins of divergent specificity and (ii) two cellulases, each bearing a dockerin complementary to one of the divergent cohesins. This approach allowed the controlled integration of desired enzymes into a multiprotein complex of predetermined stoichiometry and topology. The observed enhanced synergy on recalcitrant substrates by the bifunctional designer cellulosomes was ascribed to two major factors: substrate targeting and proximity of the two catalytic components. In the present work, the capacity of the previously described chimeric cellulosomes was amplified by developing a third divergent cohesin-dockerin device. The resultant trifunctional designer cellulosomes were assayed on homogeneous and complex substrates (microcrystalline cellulose and straw, respectively) and found to be considerably more active than the corresponding free enzyme or bifunctional systems. The results indicate that the synergy between two prominent cellulosomal enzymes (from the family-48 and -9 glycoside hydrolases) plays a crucial role during the degradation of cellulose by cellulosomes and that one dominant family-48 processive endoglucanase per complex is sufficient to achieve optimal levels of synergistic activity. Furthermore cooperation within a cellulosome chimera between cellulases and a hemicellulase from different microorganisms was achieved, leading to a trifunctional complex with enhanced activity on a complex substrate.

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Section: Discussionmentioning

confidence: 99%

Section: Selection and Characterization Of A Third Divergentmentioning

confidence: 99%

Action of Designer Cellulosomes on Homogeneous Versus Complex Substrates

Fierobe

Mingardon

Mechaly

et al. 2005

Journal of Biological Chemistry

217

View full text Add to dashboard Cite

show abstract

“…Within the cellulosome, cellulases and other glycoside hydrolases [2,3] are assembled onto multidomain scaffoldin proteins for efficient degradation of cellulosic substrates [4]. Cellulosome assembly is achieved by binding of dockerin domains from enzymes to cohesin domains in scaffoldin, and interaction with the substrate is mediated by one or more carbohydrate-binding modules (CBMs) on the scaffoldin [1,5].…”

Section: Introductionmentioning

confidence: 99%

A diverse set of family 48 bacterial glycoside hydrolase cellulases created by structure‐guided recombination

et al. 2012

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Sequence diversity within a family of functional enzymes provides a platform for elucidating structure-function relationships and for protein engineering to improve properties important for applications. Access to nature's vast sequence diversity is often limited by the fact that only a few enzymes have been characterized in a given family. Here, we recombined the catalytic domains of three glycoside hydrolase family 48 bacterial cellulases (Cel48; EC 3.2.1.176) -Clostridium cellulolyticum CelF, Clostridium stercorarium CelY, and Clostridium thermocellum CelS -to create a diverse library of Cel48 enzymes with an average of 106 mutations from the closest native enzyme. Within this set, we found large variations in properties such as the functional temperature range, stability, and specific activity on crystalline cellulose. We showed that functional status and stability were predictable from simple linear models of the sequence-property data: recombined protein fragments contributed additively to these properties in a given chimera. Using this, we correctly predicted sequences that were as stable as any of the native Cel48 enzymes described to date. The characterization of 60 active Cel48 chimeras expands the number of characterized Cel48 enzymes from 13 to 73. Our work illustrates the role that structure-guided recombination can play in helping to identify sequencefunction relationships within a family of enzymes by supplementing natural diversity with synthetic diversity.

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“…On the bacterial side, even fewer clones are available. These are limited to Cellvibrio japonicum (DeBoy et al 2008; formerly Pseudomonas fluorescens; Ferreira et al 1993), Burkholderia multivorans (Rashamuse et al 2007), Butyrivibrio fibrisolvens (Dalrymple et al 1996;Dalrymple and Swadling 1997), Pseudoalteromonas halosplanktis (Aurilia et al 2008), Clostridium thermocellum cellulosome (Blum et al 2000), and Prevotella ruminicola (Dodd et al 2009). …”

Section: Introductionmentioning

confidence: 99%