Heterologous expression and characterization of a novel branching enzyme from the thermoalkaliphilic anaerobic bacterium Anaerobranca gottschalkii

Thiemann, Volker; Saake, Bodo; Vollstedt, Annah; Schäfer, Thomas; Puls, Jürgen; Bertoldo, Costanzo; Freudl, Roland; Antranikian, Garabed

doi:10.1007/s00253-005-0248-7

Cited by 37 publications

(22 citation statements)

References 44 publications

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“…Their specific activities on amylose V were 621.3 U/mg for GBE Dg and 404.2 U/mg for GBE Dr (Table 3). These are relatively high values compared to specific activities previously reported for GH13 GBEs from E. coli, A. aeolicus, G. stearothermophilus, and A. gottschalkii (9,36,38,41). Only the R. obamensis GBE showed a comparable specific activity (33).…”

Section: Resultsmentioning

confidence: 86%

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The Unique Branching Patterns of Deinococcus Glycogen Branching Enzymes Are Determined by Their N-Terminal Domains

Palomo

Kralj

Maarel

et al. 2009

Appl Environ Microbiol

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Glycogen branching enzymes (GBE) or 1,4-␣-glucan branching enzymes (EC 2.4.1.18) introduce ␣-1,6 branching points in ␣-glucans, e.g., glycogen. To identify structural features in GBEs that determine their branching pattern specificity, the Deinococcus geothermalis and Deinococcus radiodurans GBE (GBE Dg and GBE Dr , respectively) were characterized. Compared to other GBEs described to date, these Deinococcus GBEs display unique branching patterns, both transferring relatively short side chains. In spite of their high amino acid sequence similarity (88%) the D. geothermalis enzyme had highest activity on amylose while the D. radiodurans enzyme preferred amylopectin. The side chain distributions of the products were clearly different: GBE Dg transferred a larger number of smaller side chains; specifically, DP5 chains corresponded to 10% of the total amount of transferred chains, versus 6.5% for GBE Dr . GH13-type GBEs are composed of a central (␤/␣) barrel catalytic domain and an N-terminal and a C-terminal domain. Characterization of hybrid Deinococcus GBEs revealed that the N2 modules of the N domains largely determined substrate specificity and the product branching pattern. The N2 module has recently been annotated as a carbohydrate binding module (CBM48). It appears likely that the distance between the sugar binding subsites in the active site and the CBM48 subdomain determines the average lengths of side chains transferred.

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Section: Resultsmentioning

confidence: 86%

“…These two GBEs (GBE Dg and GBE Dr , respectively) generate unique branching patterns by transferring glucosidic chains that are shorter than those of other GBEs reported to date (9,36,38,41). To investigate the role of the different domains in these enzymes, chimeras of GBE Dg and GBE Dr were constructed.…”

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confidence: 99%

The Unique Branching Patterns of Deinococcus Glycogen Branching Enzymes Are Determined by Their N-Terminal Domains

Palomo

Kralj

Maarel

et al. 2009

Appl Environ Microbiol

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“…The mature domain of Tat substrates indeed appears not to contain species-specific recognition signals for their respective Tat translocons. This is exemplified by many artificial Tat substrates which are Tat dependently translocated as a result of an N-terminal fusion to a natural Tat signal sequence, most frequently that of the E. coli TMAO reductase TorA (6,11,15,20,24,34,35,46,47).…”

Section: Discussionmentioning

confidence: 99%

Conservation and Variation between Rhodobacter capsulatus and Escherichia coli Tat Systems

Lindenstrauß

Brüser

2006

J Bacteriol

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The Tat system allows the translocation of folded and often cofactor-containing proteins across biological membranes. Here, we show by an interspecies transfer of a complete Tat translocon that Tat systems are largely, but not fully, interchangeable even between different classes of proteobacteria. The Tat apparatus from the ␣-proteobacterium Rhodobacter capsulatus was transferred to a Tat-deficient Escherichia coli strain, which is a ␥-proteobacterium. Similar to that of E. coli, the R. capsulatus Tat system consists of three components, rc-TatA, rc-TatB, and rc-TatC. A fourth gene (rc-tatF) is present in the rc-tatABCF operon which has no apparent relevance for translocation. The translational starts of rc-tatC and rc-tatF overlap in four nucleotides (ATGA) with the preceding tat genes, pointing to efficient translational coupling of rc-tatB, rc-tatC, and rc-tatF. We show by a variety of physiological and biochemical assays that the R. capsulatus Tat system functionally targets the E. coli Tat substrates TorA, AmiA, AmiC, and formate dehydrogenase. Even a Tat substrate from a third organism is accepted, demonstrating that usually Tat systems and Tat substrates from different proteobacteria are compatible with each other. Only one exceptional Tat substrate of E. coli, a membraneanchored dimethyl sulfoxide (DMSO) reductase, was not targeted by the R. capsulatus Tat system, resulting in a DMSO respiration deficiency. Although the general features of Tat substrates and translocons are similar between species, the data indicate that details in the targeting pathways can vary considerably.The twin-arginine translocation (Tat) system is known to translocate folded proteins across bacterial energy-transducing membranes (26). Three functionally distinct components of Tat systems have been identified, namely, TatA, TatB, and TatC. TatA and TatB resemble each other, and single-aminoacid exchanges can render TatA to a TatB-substituting component (6). Some organisms contain paralogs of individual components, and many organisms contain only one TatA/Blike component (13). Substrates of the Tat system are synthesized with an N-terminal signal sequence which contains the eponymous twin-arginine pattern (26). As the individual Tat substrates differ between species, it appears possible that Tat systems preferentially recognize their respective substrates. This view has been supported by studies on the transport of glucose:fructose oxidoreductase (GFOR) from Zymomonas mobilis in Escherichia coli (7). GFOR was not compatible with the Tat system from E. coli but became compatible when the signal sequence was exchanged with that from E. coli trimethylamine N-oxide (TMAO) reductase. Also, a Bacillus subtilis PhoD signal LacZ fusion was not translocated by the E. coli Tat system but became translocated by the PhoD-specific Bacillus Tat system (30). On the other hand, the Tat substrates alkaline phosphatase from Thermus thermophilus (1), highpotential iron-sulfur protein (HiPIP) from Allochromatium vinosum (9), and PlcH from Pseudomona...

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“…2). The cyclodextrin glucanotransferases (CGTases) disproportionate ␣-(1,4) bonds to yield cyclic ␣-(1,4)-oligosaccharides (cyclodextrins) while branching enzymes cleave ␣-(1,4) bonds before reattachment of the glucan chain by an ␣-(1,6) bond (MacGregor et al, 2001;Thiemann et al, 2006) (Fig. 2).…”

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confidence: 99%

Starch and α-glucan acting enzymes, modulating their properties by directed evolution

Kelly

Dijkhuizen

Leemhuis

2009

Journal of Biotechnology

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a b s t r a c tStarch is the major food reserve in plants and forms a large part of the daily calorie intake in the human diet. Industrially, starch has become a major raw material in the production of various products including bio-ethanol, coating and anti-staling agents. The complexity and diversity of these starch based industries and the demand for high quality end products through extensive starch processing, can only be met through the use of a broad range of starch and ␣-glucan modifying enzymes. The economic importance of these enzymes is such that the starch industry has grown to be the largest market for enzymes after the detergent industry. However, as the starch based industries expand and develop the demand for more efficient enzymes leading to lower production cost and higher quality products increases. This in turn stimulates interest in modifying the properties of existing starch and ␣-glucan acting enzymes through a variety of molecular evolution strategies. Within this review we examine and discuss the directed evolution strategies applied in the modulation of specific properties of starch and ␣-glucan acting enzymes and highlight the recent developments in the field of directed evolution techniques which are likely to be implemented in the future engineering of these enzymes.

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Heterologous expression and characterization of a novel branching enzyme from the thermoalkaliphilic anaerobic bacterium Anaerobranca gottschalkii

Cited by 37 publications

References 44 publications

The Unique Branching Patterns of Deinococcus Glycogen Branching Enzymes Are Determined by Their N-Terminal Domains

The Unique Branching Patterns of Deinococcus Glycogen Branching Enzymes Are Determined by Their N-Terminal Domains

Conservation and Variation between Rhodobacter capsulatus and Escherichia coli Tat Systems

Starch and α-glucan acting enzymes, modulating their properties by directed evolution

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