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Gene cloning and characterization of a vanadium-dependent bromoperoxidase from the red alga Laurencia saitoi, a producer of brominated diterpenoids and triterpenoids

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Abstract

The gene encoding vanadium-dependent bromoperoxidase (V-BPO) was cloned for the first time from the red alga Laurencia saitoi, which produces pharmaceutically promising brominated diterpenoids and triterpenoids. The molecular weight of V-BPO from L. saitoi (LsVBPO1) was the highest (77.0 kDa) among previously reported V-BPOs from Laurencia with a peptide insertion Asn194-Ser221 containing short Gln repeats. It shares approximately 60% amino acid sequence identity with V-BPOs from L. nipponica (LnVBPO1 and LnVBPO2) and L. okamurae (LoVBPO1a and LoVBPO2a). Heterologously expressed LsVBPO1 in Escherichia coli was partially purified and exhibited low but significant bromination activity of 38 U mg−1 protein using monochlorodimedone. The pH optimum was 8.0, which was more alkaline than that for LnVBPOs and LoVBPO2a (pH 7.0). The Km for H2O2 was 0.04 mM, comparable to LnVBPO1 (0.026 mM), LnVBPO2 (0.025 mM), and LoVBPO2a (0.014 mM). LsVPBO1 retained its bromination activity until 45 °C for 20 min. When incubated at 55 °C for 20 min, catalytic activity decreased rapidly, as shown for LnVBPO1 and LoVBPO2a (retained at 45 °C, decreased at 55 °C) and LnVBPO2 (retained at 55 °C, decreased at 65 °C). Unlike other V-BPOs from Laurencia (LnVBPO1, LnVBPO2, and LoVBPO2a), dialysis and concentration during purification process were rapidly inactivated LsVBPO1, suggesting its structural instability.

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Data availability

Raw data were generated at Hokkaido University. Data supporting the findings of this study are available from the corresponding authors K.K. and T.O. on request.

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Acknowledgements

Kensuke Kaneko thanks the GCOE program of the Graduate School of Environmental Science at Hokkaido University. We thank Prof. Minoru Suzuki and Dr. Tsuyoshi Abe (Hokkaido University) for their valuable advice and suggestions. We are grateful to the staff of the Oshoro Marine Station, Field Science Center for Northern Biosphere, Hokkaido University, for their support and assistance during the field survey. We would like to thank Editage (www.editage.com) for English language editing.

Funding

This work was supported by a JSPS KAKENHI grant (No. 16H04975).

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Authors and Affiliations

  1. Graduate School of Environmental Earth Science, Hokkaido University, Sapporo, 060-0810, Japan

    Kensuke Kaneko, Daiki Kobayashi, Shiro Masaki, Kenji Washio, Masaaki Morikawa & Tatsufumi Okino

  2. Hachinohe College of Technology, Aomori, 039-1192, Japan

    Kensuke Kaneko

  3. Faculty of Environmental Earth Science, Hokkaido University, Sapporo, 060-0810, Japan

    Kenji Washio, Masaaki Morikawa & Tatsufumi Okino

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Contributions

Kensuke Kaneko conceived and designed experiments, conducted all experiments, and wrote the paper. Daiki Kobayashi, Shiro Masaki, and Kenji Washio cloned and heterologously expressed LsVBPO1. Masaaki Morikawa wrote and revised the manuscript accordingly. Tatsufumi Okino conceived and designed experiments, and wrote the manuscript. All authors have reviewed the manuscript.

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Correspondence to Kensuke Kaneko or Tatsufumi Okino.

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Kaneko, K., Kobayashi, D., Masaki, S. et al. Gene cloning and characterization of a vanadium-dependent bromoperoxidase from the red alga Laurencia saitoi, a producer of brominated diterpenoids and triterpenoids. J Appl Phycol 35, 1443–1452 (2023). https://doi.org/10.1007/s10811-023-02953-w

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