Isolation and Characterization of Genes Responsible for Naphthalene Degradation from Thermophilic Naphthalene Degrader, Geobacillus sp. JF8

Daisuke Miyazawa; Le Thi Ha Thanh; Akio Tani; Masaki Shintani; Nguyen Hoang Loc; Takashi Hatta; Kazuhide Kimbara

doi:10.3390/microorganisms8010044

Isolation and Characterization of Genes Responsible for Naphthalene Degradation from Thermophilic Naphthalene Degrader, Geobacillus sp. JF8

Microorganisms. 2019 Dec 24;8(1):44. doi: 10.3390/microorganisms8010044.

Authors

Daisuke Miyazawa¹, Le Thi Ha Thanh^{2

3}, Akio Tani¹, Masaki Shintani^{4

5

6}, Nguyen Hoang Loc³, Takashi Hatta⁷, Kazuhide Kimbara^{2

5}

Affiliations

¹ Institute of Plant Science and Resources, Okayama University, 2-20-1 Chuo, Kurashiki, Okayama 710-0046, Japan.
² Department of Environment and Energy System, Graduate School of Science and Technology, Shizuoka University, 3-5-1 Johoku, Naka-ku, Hamamatsu, Shizuoka 432-8011, Japan.
³ Institute of Bioactive Compounds, University of Sciences, Hue University, Hue, Thua Thien Hue 530000, Vietnam.
⁴ Department of Bioscience, Graduate School of Science and Technology, Shizuoka University, 3-5-1 Johoku, Naka-ku, Hamamatsu, Shizuoka 432-8561, Japan.
⁵ Department of Engineering, Graduate School of Integrated Science and Technology, Shizuoka University, 3-5-1 Johoku, Naka-ku, Hamamatsu, Shizuoka 432-8561, Japan.
⁶ Research Institute of Green Science and Technology, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, Shizuoka 422-8529, Japan.
⁷ Department of Biomedical Engineering, Okayama University of Science, 1-1 Ridai-cho, Kita-ku, Okayama 703-8232, Japan.

Abstract

Geobacillus sp. JF8 is a thermophilic biphenyl and naphthalene degrader. To identify the naphthalene degradation genes, cis-naphthalene dihydrodiol dehydrogenase was purified from naphthalene-grown cells, and its N-terminal amino acid sequence was determined. Using a DNA probe encoding the N-terminal region of the dehydrogenase, a 10-kb DNA fragment was isolated. Upstream of nahB, a gene for dehydrogenase, there were two open reading frames which were designated as nahAc and nahAd, respectively. The products of nahAc and nahAd were predicted to be alpha and beta subunit of ring-hydroxylating dioxygenases, respectively. Phylogenetic analysis of amino acid sequences of NahB indicated that it did not belong to the cis-dihydrodiol dehydrogenase group that includes those of classical naphthalene degradation pathways. Downstream of nahB, four open reading frames were found, and their products were predicted as meta-cleavage product hydrolase, monooxygenase, dehydrogenase, and gentisate 1,2-dioxygenase, respectively. A reverse transcriptase-PCR analysis showed that transcription of nahAcAd was induced by naphthalene. These findings indicate that we successfully identified genes involved in the upper pathway of naphthalene degradation from a thermophilic bacterium.

Keywords: Geobacillus; naphthalene; thermophile.