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Cloning and Sequencing of Alginate Lyase Genes from Deep-Sea Strains of Vibrio and Agarivorans and Characterization of a New Vibrio Enzyme

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Abstract

Four alginate lyase genes were cloned and sequenced from the genomic DNAs of deep-sea bacteria, namely members of Vibrio and Agarivorans. Three of them were from Vibrio sp. JAM-A9m, which encoded alginate lyases, A9mT, A9mC, and A9mL. A9mT was composed of 286 amino acids and 57% homologous to AlxM of Photobacterium sp. A9mC (221 amino acids) and A9mL (522 amino acids) had the highest degree of similarity to two individual alginate lyases of Vibrio splendidus with 74% and 84% identity, respectively. The other gene for alginate lyase, A1mU, was shotgun cloned from Agarivorans sp. JAM-A1m. A1mU (286 amino acids) showed the highest homology to AlyVOA of Vibrio sp. with 76% identity. All alginate lyases belong to polysaccharide lyase family 7, although, they do not show significant similarity to one another with 14% to 58% identity. Among the above lyases, the recombinant A9mT was purified to homogeneity and characterized. The molecular mass of A9mT was around 28 kDa. The enzyme was remarkably salt activated and showed the highest thermal stability in the presence of NaCl. A9mT favorably degraded mannuronate polymer in alginate. We discussed substrate specificities of family 7 alginate lyases based on their conserved amino acid sequences.

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Acknowledgments

We appreciate the captain and crew of the R/V Natushima and the ROV Hyper Dolphin operation team for their technical support in sampling. We also thank the chief scientist, Prof. K. Kubokawa of the Ocean Research Institute, University of Tokyo, and scientists aboard the NT05-12 cruise.

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Authors and Affiliations

  1. Institute of Biogeoscience, Japan Agency for Marine-Earth Science and Technology (JAMSTEC), 2-15 Natsushima, Yokosuka, Japan, 237-0061

    Kohsuke Uchimura, Masayuki Miyazaki, Yuichi Nogi, Tohru Kobayashi & Koki Horikoshi

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  1. Kohsuke Uchimura
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  2. Masayuki Miyazaki
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  3. Yuichi Nogi
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  4. Tohru Kobayashi
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  5. Koki Horikoshi
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Corresponding author

Correspondence to Tohru Kobayashi.

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Supplementary Table

Amino acid among alginate lyases of strains A9m amd A1m (GIF 75 kb)

Supplementary Table

High resolution image (TIFF 55 kb)

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Uchimura, K., Miyazaki, M., Nogi, Y. et al. Cloning and Sequencing of Alginate Lyase Genes from Deep-Sea Strains of Vibrio and Agarivorans and Characterization of a New Vibrio Enzyme. Mar Biotechnol 12, 526–533 (2010). https://doi.org/10.1007/s10126-009-9237-7

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  • DOI: https://doi.org/10.1007/s10126-009-9237-7

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