Purification and properties of cellobiose phosphorylase from Clostridium thermocellum

Tanaka, Koujirou; Kawaguchi, Takashi; Imada, Yukio; Ooi, Toshihiko; Arai, Motoo

doi:10.1016/0922-338x(95)90605-y

Cited by 21 publications

(21 citation statements)

References 17 publications

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“…Because the phosphorylation uses inorganic phosphate as a donor, the phosphorolytic mechanism is an ATP-saving mechanism more often associated with some cellulolytic bacteria (5,12,15,20). All known cellobiose phosphorylases are cytoplasmic, as they lack signal peptides, and experimental data support this notion (1,19).…”

supporting

confidence: 54%

Engineering Escherichia coli Cells for Cellobiose Assimilation through a Phosphorolytic Mechanism

Sekar

Shin

Chen

2012

Appl Environ Microbiol

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We report here the first engineering effort for Escherichia coli biocatalysts to assimilate cellobiose through a phosphorolytic mechanism. Cytoplasmic expression of the Saccharophagus cellobiose phosphorylase was shown to enable E. coli to use cellobiose. Subsequent knockout and complementation studies provided solid evidence that the endogenous LacY was responsible for the transport of cellobiose.

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supporting

confidence: 54%

Engineering Escherichia coli Cells for Cellobiose Assimilation through a Phosphorolytic Mechanism

Sekar

Shin

Chen

2012

Appl Environ Microbiol

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“…CbP is widely distributed in cellobiose-utilizing bacteria, both anaerobic (9,10,23,249,667,738,740) and aerobic (600), and the enzyme has even been found in the hyperthermophile Thermotoga neopolitana (769). The CbP from Cellvibrio gilvus is particularly interesting in that its wide glucosyl acceptor specificity has permitted the in vitro synthesis of numerous unusual di-and trisaccharides (670).…”

Section: Physiology Of Cellulolytic Microorganismsmentioning

confidence: 99%

Microbial Cellulose Utilization: Fundamentals and Biotechnology

Lynd

Weimer

Zyl³

et al. 2002

Microbiol Mol Biol Rev

4,006

2,868

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Fundamental features of microbial cellulose utilization are examined at successively higher levels of aggregation encompassing the structure and composition of cellulosic biomass, taxonomic diversity, cellulase enzyme systems, molecular biology of cellulase enzymes, physiology of cellulolytic microorganisms, ecological aspects of cellulase-degrading communities, and rate-limiting factors in nature. The methodological basis for studying microbial cellulose utilization is considered relative to quantification of cells and enzymes in the presence of solid substrates as well as apparatus and analysis for cellulose-grown continuous cultures. Quantitative description of cellulose hydrolysis is addressed with respect to adsorption of cellulase enzymes, rates of enzymatic hydrolysis, bioenergetics of microbial cellulose utilization, kinetics of microbial cellulose utilization, and contrasting features compared to soluble substrate kinetics. A biological perspective on processing cellulosic biomass is presented, including features of pretreated substrates and alternative process configurations. Organism development is considered for “consolidated bioprocessing” (CBP), in which the production of cellulolytic enzymes, hydrolysis of biomass, and fermentation of resulting sugars to desired products occur in one step. Two organism development strategies for CBP are examined: (i) improve product yield and tolerance in microorganisms able to utilize cellulose, or (ii) express a heterologous system for cellulose hydrolysis and utilization in microorganisms that exhibit high product yield and tolerance. A concluding discussion identifies unresolved issues pertaining to microbial cellulose utilization, suggests approaches by which such issues might be resolved, and contrasts a microbially oriented cellulose hydrolysis paradigm to the more conventional enzymatically oriented paradigm in both fundamental and applied contexts

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“…C. thermocellum was described as a cellulose-hydrolysing bacterium, which grows on cellulose and cellodextrins, but not on pentoses and moreover, does not even readily grow on glucose. It was discussed that the uptake of oligocellodextrins is energetically favourable, because (1) energy is consumed for only one transportation event for more glucose residues, and (2) oligodextrins are degraded by intracellular cellobiose-and cellodextrin-phosphorylases producing glucose 1-phosphate (Tanaka et al, 1995;Reichenbecher et al, 1997;Lynd et al, 2002). It is not known if cellobiose or cellodextrin phosphorylases hydrolyse b-1,3-oligodextrins.…”

Section: Discussionmentioning

confidence: 99%

Lic16A of Clostridium thermocellum, a non-cellulosomal, highly complex endo-β-1,3-glucanase bound to the outer cell surface

Fuchs¹,

Zverlov

Velikodvorskaya

et al. 2003

Microbiology

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Clostridium thermocellum produces one major b-1,3-glucanase. Genomic DNA fragments containing the gene were cloned from two strains, DSM1237T (6848 bp) and F7 (9766 bp).Overlapping sequences were 99?9 % identical. The nucleotide sequences contained reading frames for a putative transposase, endo-b-1,3-1,4-glucanase CelC, a putative transcription regulator of the LacI type, b-1,3-glucanase Lic16A and a putative membrane protein. The licA genes of both strains encoded an identical protein of 1324 aa with a calculated molecular mass of 148 kDa. Lic16A is an unusually complex protein consisting of a leader peptide, a threefold repeat of an S-layer homologous module (SLH), an unknown module, a catalytic module of glycosyl hydrolase family 16 and a fourfold repeat of a carbohydrate-binding module of family CBM4a. The recombinant Lic16A protein was characterized as an endo-1,3(4)-b-glucanase with a specific activity of 2680 and 340 U mg 21 and a K m of 0?94 and 2?1 mg ml 21 towards barley b-glucan and laminarin, respectively. It was specific for b-glucans containing b-1,3-linkages with an optimum temperature of 70˚C at pH 6?0. The N-terminal SLH modules were cleaved from the protein as well in Escherichia coli as in C. thermocellum, but nevertheless bound tightly to the rest of the protein. Lic16A was located on the cell surface from which it could be purified after fractionated solubilization. Its inducible production allowed C. thermocellum to grow on b-1,3-or b-1,3-1,4-glucan.

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Purification and properties of cellobiose phosphorylase from Clostridium thermocellum

Cited by 21 publications

References 17 publications

Engineering Escherichia coli Cells for Cellobiose Assimilation through a Phosphorolytic Mechanism

Engineering Escherichia coli Cells for Cellobiose Assimilation through a Phosphorolytic Mechanism

Microbial Cellulose Utilization: Fundamentals and Biotechnology

Lic16A of Clostridium thermocellum, a non-cellulosomal, highly complex endo-β-1,3-glucanase bound to the outer cell surface

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