Monomer sequence and acetylation pattern in some bacterial alginates

Skj»k-Bræk, Gudmund; Grasdalen, Hans; Lársen, Bjørn

doi:10.1016/s0008-6215(00)90036-3

Cited by 287 publications

(191 citation statements)

References 21 publications

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“…These data show that, at least under the growth conditions used here, A. vinelandii AlgG contributes very little to epimerization. In comparison, Pseudomonas AlgG normally introduces more than 20% G in its alginates (21,54). It seems possible that the A. vinelandii AlgG activity has become reduced in evolution due to the development of key roles for the AlgE MEs.…”

Section: Resultsmentioning

confidence: 99%

“…In contrast, the fractional contents and sequence distributions of the M and G residues vary widely (54), and alginate structures are described as being composed of G blocks, M blocks, and MG blocks of various lengths. G blocks are of great biological and applied significance since they are a prerequisite for the formation of strong polymer gels in the presence of divalent cations like Ca 2ϩ (26).…”

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confidence: 99%

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Identification and Characterization of anAzotobacter vinelandiiType I Secretion System Responsible for Export of the AlgE-Type Mannuronan C-5-Epimerases

et al. 2006

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Alginate is a linear copolymer of ␤-D-mannuronic acid and its C-5-epimer, ␣-L-guluronic acid. During biosynthesis, the polymer is first made as mannuronan, and various fractions of the monomers are then epimerized to guluronic acid by mannuronan C-5-epimerases. The Azotobacter vinelandii genome encodes a family of seven extracellular such epimerases (AlgE1 to AlgE7) which display motifs characteristic for proteins secreted via a type I pathway. Putative ATPase-binding cassette regions from the genome draft sequence of the A. vinelandii OP strain and experimentally verified type I transporters from other species were compared. This analysis led to the identification of one putative A. vinelandii type I system (eexDEF). The corresponding genes were individually disrupted in A. vinelandii strain E, and Western blot analysis using polyclonal antibodies against all AlgE epimerases showed that these proteins were present in wild-type culture supernatants but absent from the eex mutant supernatants. Consistent with this, the wild-type strain and the eex mutants produced alginate with about 20% guluronic acid and almost pure mannuronan (<2% guluronic acid), respectively. The A. vinelandii wild type is able to enter a particular desiccation-tolerant resting stage designated cyst. At this stage, the cells are surrounded by a rigid coat in which alginate is a major constituent. Such a coat was formed by wild-type cells in a particular growth medium but was missing in the eex mutants. These mutants were also found to be unable to survive desiccation. The reason for this is probably that continuous stretches of guluronic acid residues are needed for alginate gel formation to take place.

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

Identification and Characterization of anAzotobacter vinelandiiType I Secretion System Responsible for Export of the AlgE-Type Mannuronan C-5-Epimerases

et al. 2006

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“…Acetylation of carbohydrates is not restricted to land plants and also has been observed in fungi (Cryptococcus neoformans; glucoronoxylomannan [Cherniak and Sundstrom, 1994]), mammals (sialic acids [Hutchins et al, 1988]), and bacteria (peptidoglycan [Clarke and Dupont, 1992]; alginate [Skjåk-Braek et al, 1986], and cellulose [Spiers et al, 2003]). These nonplant organisms contain acetylation machinery that has homology to the RWA and TBL proteins, sometimes as parts of a single polypeptide (Gille and Pauly, 2012).…”

Section: Involvement Of Axy9 In the Polysaccharide Acetylation Pathwaymentioning

confidence: 99%

The Role of the Plant-Specific ALTERED XYLOGLUCAN9 Protein in Arabidopsis Cell Wall PolysaccharideO-Acetylation

et al. 2015

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A mutation in the ALTERED XYLOGLUCAN9 (AXY9) gene was found to be causative for the decreased xyloglucan acetylation phenotype of the axy9.1 mutant, which was identified in a forward genetic screen for Arabidopsis (Arabidopsis thaliana) mutants. The axy9.1 mutant also exhibits decreased O-acetylation of xylan, implying that the AXY9 protein has a broad role in polysaccharide acetylation. An axy9 insertional mutant exhibits severe growth defects and collapsed xylem, demonstrating the importance of wall polysaccharide O-acetylation for normal plant growth and development. Localization and topological experiments indicate that the active site of the AXY9 protein resides within the Golgi lumen. The AXY9 protein appears to be a component of the plant cell wall polysaccharide acetylation pathway, which also includes the REDUCED WALL ACETYLATION and TRICHOME BIREFRINGENCE-LIKE proteins. The AXY9 protein is distinct from the TRICHOME BIREFRINGENCE-LIKE proteins, reported to be polysaccharide acetyltransferases, but does share homology with them and other acetyltransferases, suggesting that the AXY9 protein may act to produce an acetylated intermediate that is part of the O-acetylation pathway.

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“…Biosynthesis of alginate involves the initial production of polymannuronate followed by the introduction of G residues catalyzed by mannuronan C-5 epimerases (5-7). Pseudomonas alginates differ from those obtained by other sources in lacking G-blocks, and bacterial alginates, as opposed to the seaweed polymers, can be acetylated to various degrees at positions O-2 and/or O-3 on the M residues (8). The material properties of alginates are determined by intrinsic properties like the polymer chain length, acetylation level, and the monomer composition and distribution pattern.…”

mentioning

confidence: 99%

“…Hence there is an emerging need for more detailed information concerning the alginate fine structure. Although the total monomer composition and the dyad and triad frequencies can be elucidated by, for example, high resolution NMR spectroscopy (8), there is limited knowledge on the distribution and the absolute length of the various block types.…”

mentioning

confidence: 99%

Isolation of Mutant Alginate Lyases with Cleavage Specificity for Di-guluronic Acid Linkages

Tøndervik

Klinkenberg

Aarstad

et al. 2010

Journal of Biological Chemistry

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Alginates are commercially valuable and complex polysaccharides composed of varying amounts and distribution patterns of 1-4-linked ␤-D-mannuronic acid (M) and ␣-L-guluronic acid (G). This structural variability strongly affects polymer physicochemical properties and thereby both commercial applications and biological functions. One promising approach to alginate fine structure elucidation involves the use of alginate lyases, which degrade the polysaccharide by cleaving the glycosidic linkages through a ␤-elimination reaction. For such studies one would ideally like to have different lyases, each of which cleaves only one of the four possible linkages in alginates: G-G, G-M, M-G, and M-M. So far no lyase specific for only G-G linkages has been described, and here we report the construction of such an enzyme by mutating the gene encoding Klebsiella pneumoniae lyase AlyA (a polysaccharide lyase family 7 lyase), which cleaves both G-G and G-M linkages. After error-prone PCR mutagenesis and high throughput screening of ϳ7000 lyase mutants, enzyme variants with a strongly improved G-G specificity were identified. Furthermore, in the absence of Ca 2؉ , one of these lyases (AlyA5) was found to display no detectable activity against G-M linkages. G-G linkages were cleaved with ϳ10% of the optimal activity under the same conditions. The substitutions conferring altered specificity to the mutant enzymes are located in conserved regions in the polysaccharide lyase family 7 alginate lyases. Structure-function analyses by comparison with the known three-dimensional structure of Sphingomonas sp. A1 lyase A1-II suggests that the improved G-G specificity might be caused by increased affinity for nonproductive binding of the alternating G-M structure.

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Monomer sequence and acetylation pattern in some bacterial alginates

Cited by 287 publications

References 21 publications

Identification and Characterization of anAzotobacter vinelandiiType I Secretion System Responsible for Export of the AlgE-Type Mannuronan C-5-Epimerases

Identification and Characterization of anAzotobacter vinelandiiType I Secretion System Responsible for Export of the AlgE-Type Mannuronan C-5-Epimerases

The Role of the Plant-Specific ALTERED XYLOGLUCAN9 Protein in Arabidopsis Cell Wall PolysaccharideO-Acetylation

Isolation of Mutant Alginate Lyases with Cleavage Specificity for Di-guluronic Acid Linkages

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