The roles of oxygen and alginate-lyase in determining the molecular weight of alginate produced by Azotobacter vinelandii

2004 ◽  
Vol 63 (6) ◽  
pp. 742-747 ◽  
Author(s):  
M. A. Trujillo-Rold�n ◽  
S. Moreno ◽  
G. Esp�n ◽  
E. Galindo
Keyword(s):  
Molecular Weight ◽  
Azotobacter Vinelandii ◽  
Alginate Lyase

scholarly journals Mechanistic basis for understanding the dual activities of the bifunctional Azotobacter vinelandii mannuronan C-5 epimerase and alginate lyase AlgE7

2021 ◽  
Author(s):  
Margrethe Gaardløs ◽  
Tonje Marita Bjerkan Heggeset ◽  
Anne Tøndervik ◽  
David Tezé ◽  
Birte Svensson ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Functional Properties ◽  
Catalytic Mechanism ◽  
Azotobacter Vinelandii ◽  
Brown Seaweed ◽  
Alginate Lyase ◽  
High Sequence Identity ◽  
Monomer Composition ◽  
Lyase Activity ◽  
Mechanistic Basis

The structure and functional properties of alginates are dictated by the monomer composition and molecular weight distribution. Mannuronan C-5 epimerases determine the monomer composition by catalysing the epimerization of β- d -mannuronic acid residues (M) into α- l -guluronic acid residues (G). The molecular weight is affected by alginate lyases, which catalyse a β-elimination mechanism that cleaves alginate chains. The reaction mechanisms for the epimerization and lyase reactions are similar and some enzymes can perform both reactions. These dualistic enzymes share high sequence identity with mannuronan C-5 epimerases without lyase activity. The mechanism behind their activity and the amino acid residues responsible for it are still unknown. We investigate mechanistic determinants involved in the bifunctional epimerase and lyase activity of AlgE7 from Azotobacter vinelandii . Based on sequence analyses, a range of AlgE7 variants were constructed and subjected to activity assays and product characterization by NMR. Our results show that calcium promotes lyase activity whereas NaCl reduces the lyase activity of AlgE7. By using defined poly-M and poly-MG substrates, the preferred cleavage sites of AlgE7 were found to be M|XM and G|XM, where X can be either M or G. From the study of AlgE7 mutants, R148 was identified as an important residue for the lyase activity, and the point mutant R148G resulted in an enzyme with only epimerase activity. Based on the results obtained in the present study we suggest a unified catalytic reaction mechanism for both epimerase and lyase activity where H154 functions as the catalytic base and Y149 as the catalytic acid. Importance Post-harvest valorisation and upgrading of algal constituents is a promising strategy in the development of a sustainable bioeconomy based on algal biomass. In this respect, alginate epimerases and lyases are valuable enzymes for tailoring of the functional properties of alginate, a polysaccharide extracted from brown seaweed with numerous applications in food, medicine, and material industries. By providing a better understanding of the catalytic mechanism and of how the two enzyme actions can be altered by changes in reaction conditions, this study opens for further applications of bacterial epimerases and lyases in enzymatic tailoring of alginate polymers.

scholarly journals Macrophage-stimulating activities of a novel low molecular weight saccharide fragment prepared from ascophyllan with alginate lyase

2020 ◽  
Vol 67 ◽  
pp. 103839
Author(s):  
Zedong Jiang ◽  
Gang Yu ◽  
Qingyun Bao ◽  
Xu Xu ◽  
Yanbing Zhu ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Alginate Lyase ◽  
Low Molecular Weight

scholarly journals An Alginate Lyase from Azotobacter vinelandii Phage

1977 ◽  
Vol 98 (1) ◽  
pp. 223-229 ◽  
Author(s):  
I. W. Davidson ◽  
C. J. Lawson ◽  
I. W. Sutherland
Keyword(s):  
Azotobacter Vinelandii ◽  
Alginate Lyase

Partial repression of siderophore-mediated iron transport in Azotobacter vinelandii grown with mineral iron

1988 ◽  
Vol 34 (5) ◽  
pp. 675-679 ◽  
Author(s):  
William J. Page ◽  
Gregory A. Grant
Keyword(s):  
Molecular Weight ◽  
Outer Membrane ◽  
Iron Transport ◽  
Mineral Content ◽  
Azotobacter Vinelandii ◽  
Outer Membrane Proteins ◽  
Additional Component ◽  
Transport Activity ◽  
Fe Uptake ◽  
Mineral Concentrations

Azotobacter vinelandii grown in iron-limited medium containing increased amounts of the minerals olivine or glauconite produced decreased amounts of siderophores and iron-repressible outer membrane proteins. These minerals caused a relatively rapid repression of the pyoverdin-type siderophore, azotobactin, and a slower repression of the catechol siderophores, azotochelin and aminochelin. A 77 000 molecular weight outer membrane protein also was repressed with increased mineral content of the medium, but coordinate repression with any one siderophore was not evident. Cells grown with increased mineral concentrations progressively lost siderophore-mediated iron-transport activity. This loss in activity had the greatest effect on azotobactin-mediated 55Fe uptake, but catechol siderophore mediated 55Fe uptake also was depressed. These results suggested that an additional component was required for maximal iron-transport activity promoted by all the siderophores of A. vinelandii.

scholarly journals Cloning and Characterization of a Novel Alginate Lyase from Paenibacillus sp. LJ-23

Marine Drugs ◽  
2022 ◽  
Vol 20 (1) ◽  
pp. 66
Author(s):  
Mingpeng Wang ◽  
Lei Chen ◽  
Zhengyu Lou ◽  
Xueting Yuan ◽  
Guiping Pan ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Water Solubility ◽  
Degradation Products ◽  
Three Dimensional ◽  
Alginate Lyase ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Alginate Oligosaccharides ◽  
Paenibacillus Sp ◽  
Comprehensive Health

As a low molecular weight alginate, alginate oligosaccharides (AOS) exhibit improved water solubility, better bioavailability, and comprehensive health benefits. In addition, their biocompatibility, biodegradability, non-toxicity, non-immunogenicity, and gelling capability make them an excellent biomaterial with a dual curative effect when applied in a drug delivery system. In this paper, a novel alginate lyase, Algpt, was cloned and characterized from a marine bacterium, Paenibacillus sp. LJ-23. The purified enzyme was composed of 387 amino acid residues, and had a molecular weight of 42.8 kDa. The optimal pH of Algpt was 7.0 and the optimal temperature was 45 °C. The analysis of the conserved domain and the prediction of the three-dimensional structure indicated that Algpt was a novel alginate lyase. The dominant degradation products of Algpt on alginate were AOS dimer to octamer, depending on the incubation time, which demonstrated that Algpt degraded alginate in an endolytic manner. In addition, Algpt was a salt-independent and thermo-tolerant alginate lyase. Its high stability and wide adaptability endow Algpt with great application potential for the efficient preparation of AOS with different sizes and AOS-based products.

scholarly journals Cloning and Expression of the algL Gene, Encoding the Azotobacter chroococcum Alginate Lyase: Purification and Characterization of the Enzyme

1999 ◽  
Vol 181 (5) ◽  
pp. 1409-1414 ◽  
Author(s):  
Ana Peciña ◽  
Alberto Pascual ◽  
Antonio Paneque
Keyword(s):  
Gene Sequence ◽  
Azotobacter Vinelandii ◽  
Alginate Lyase ◽  
Azotobacter Chroococcum ◽  
Open Reading Frame ◽  
Cloning And Expression ◽  
Gene Encoding ◽  
Reading Frame ◽  
Encoding Gene

ABSTRACT The alginate lyase-encoding gene (algL) ofAzotobacter chroococcum was localized to a 3.1-kbEcoRI DNA fragment that revealed an open reading frame of 1,116 bp. This open reading frame encodes a protein of 42.98 kDa, in agreement with the value previously reported by us for this protein. The deduced protein has a potential N-terminal signal peptide that is consistent with its proposed periplasmic location. The analysis of the deduced amino acid sequence indicated that the gene sequence has a high homology (90% identity) to the Azotobacter vinelandii gene sequence, which has very recently been deposited in the GenBank database, and that it has 64% identity to the Pseudomonas aeruginosa gene sequence but that it has rather low homology (15 to 22% identity) to the gene sequences encoding alginate lyase in other bacteria. The A. chroococcum AlgL protein was overproduced in Escherichia coli and purified to electrophoretic homogeneity in a two-step chromatography procedure on hydroxyapatite and phenyl-Sepharose. The kinetic and molecular parameters of the recombinant alginate lyase are similar to those found for the native enzyme.

scholarly journals Enzyme-Assisted Fucoidan Extraction from Brown Macroalgae Fucus distichus subsp. evanescens and Saccharina latissima

Marine Drugs ◽  
2020 ◽  
Vol 18 (6) ◽  
pp. 296 ◽  
Author(s):  
Thuan Thi Nguyen ◽  
Maria Dalgaard Mikkelsen ◽  
Vy Ha Nguyen Tran ◽  
Vo Thi Dieu Trang ◽  
Nanna Rhein-Knudsen ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Alginate Lyase ◽  
Saccharina Latissima ◽  
Brown Macroalgae ◽  
Brown Seaweeds ◽  
Molecular Weights ◽  
Fucus Distichus ◽  
Assisted Extraction ◽  
High Yields ◽  
Enzyme Assisted Extraction

Fucoidans from brown macroalgae (brown seaweeds) have different structures and many interesting bioactivities. Fucoidans are classically extracted from brown seaweeds by hot acidic extraction. Here, we report a new targeted enzyme-assisted methodology for fucoidan extraction from brown seaweeds. This enzyme-assisted extraction protocol involves a one-step combined use of a commercial cellulase preparation (Cellic®CTec2) and an alginate lyase from Sphingomonas sp. (SALy), reaction at pH 6.0, 40 °C, removal of non-fucoidan polysaccharides by Ca2+ precipitation, and ethanol-precipitation of crude fucoidan. The workability of this method is demonstrated for fucoidan extraction from Fucus distichus subsp. evanescens (basionym Fucus evanescens) and Saccharina latissima as compared with mild acidic extraction. The crude fucoidans resulting directly from the enzyme-assisted method contained considerable amounts of low molecular weight alginate, but this residual alginate was effectively removed by an additional ion-exchange chromatographic step to yield pure fucoidans (as confirmed by 1H NMR). The fucoidan yields that were obtained by the enzymatic method were comparable to the chemically extracted yields for both F. evanescens and S. latissima, but the molecular sizes of the fucoidans were significantly larger with enzyme-assisted extraction. The molecular weight distribution of the fucoidan fractions was 400 to 800 kDa for F. evanescens and 300 to 800 kDa for S. latissima, whereas the molecular weights of the corresponding chemically extracted fucoidans from these seaweeds were 10–100 kDa and 50–100 kDa, respectively. Enzyme-assisted extraction represents a new gentle strategy for fucoidan extraction and it provides new opportunities for obtaining high yields of native fucoidan structures from brown macroalgae.

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