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Cloning, Expression, and Characterization of a Novel l-Arabinose Isomerase from the Psychrotolerant Bacterium Pseudoalteromonas haloplanktis

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Abstract

l-Arabinose isomerase (L-AI, EC 5.3.1.4) catalyzes the isomerization between l-arabinose and l-ribulose, and most of the reported ones can also catalyze d-galactose to d-tagatose, except Bacillus subtilis L-AI. In this article, the L-AI from the psychrotolerant bacterium Pseudoalteromonas haloplanktis ATCC 14393 was characterized. The enzyme showed no substrate specificity toward d-galactose, which was similar to B. subtilis L-AI but distinguished from other reported L-AIs. The araA gene encoding the P. haloplanktis L-AI was cloned and overexpressed in E. coli BL21 (DE3). The recombinant enzyme was purified by one-step nickel affinity chromatography . The enzyme displayed the maximal activity at 40 °C and pH 8.0, and showed more than 75 % of maximal activity from pH 7.5–9.0. Metal ion Mn2+ was required as optimum metal cofactor for activity simulation, but it did not play a significant role in thermostability improvement as reported previously. The Michaelis–Menten constant (K m), turnover number (k cat), and catalytic efficiency (k cat/K m) for substrate l-arabinose were measured to be 111.68 mM, 773.30/min, and 6.92/mM/min, respectively. The molecular docking results showed that the active site residues of P. haloplanktis L-AI could only immobilize l-arabinose and recognized it as substrate for isomerization.

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Acknowledgments

This work was supported by the NSFC Project (No. 21276001), the 863 Project (No. 2013AA102102), the Support Project of Jiangsu Province (No. BK20130001 and 2015-SWYY-009), and the project of Outstanding Scientific and Technological Innovation Group of Jiangsu Province (**g Wu).

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Authors and Affiliations

  1. State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, 214122, Jiangsu, China

    Wei Xu, Chen Fan, Tao Zhang, Bo Jiang & Wanmeng Mu

  2. Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi, 214122, China

    Tao Zhang, Bo Jiang & Wanmeng Mu

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  1. Wei Xu
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  2. Chen Fan
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  3. Tao Zhang
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Correspondence to Wanmeng Mu.

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Xu, W., Fan, C., Zhang, T. et al. Cloning, Expression, and Characterization of a Novel l-Arabinose Isomerase from the Psychrotolerant Bacterium Pseudoalteromonas haloplanktis . Mol Biotechnol 58, 695–706 (2016). https://doi.org/10.1007/s12033-016-9969-3

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