Complete Genome Sequence of Flavisolibacter Ginsenosidimutans T Gsoil 636 , a Ginsenoside-Converting Bacterium, Isolated from Soil Used for Cultivating Ginseng

Complete genome sequence of Flavisolibacter ginsenosidimutans T Gsoil 636 , a ginsenoside-converting bacterium, isolated from soil used for cultivating ginseng

1 2 2 1 1,3 Dong-Ho Keum , Byoung Hee Lee , Ki-Eun Lee , Soon Youl Lee , and Wan-Taek Im * 1 Department of Biotechnology, Hankyong National University, Gyeonggi-do 17579, Republic of Korea 2 Microorganism Resources Division, National Institute of Biological Resources, Incheon 22689, Republic of Korea 3 AceEMzyme Co., Ltd., Academic Industry Cooperation, Gyeonggi-do 17579, Republic of Korea

인삼 재배 토양에서 분리한 진세노사이드 전환능력이 있는 T Flavisolibacter ginsenosidimutans Gsoil 636 의 유전체 서열 분석

금동호1 ･ 이병희2 ･ 이기은2 ･ 이순열1 ･ 임완택1,3* 1 2 3 국립한경대학교 농업생명과학대학 생명공학과, 국림생물자원관 미생물자원과, (주)에이스엠자임

(Received October 30, 2019; Revised December 9, 2019; Accepted December 9, 2019)

T A yellow-colored, circular, convex, rod-shaped baterial strain Im, 2007). Strain Gsoil 636 is Gram-negative-bacterium, rod- T designated Flavisolibacter ginsenosidimutans Gsoil 636 was shaped, non-motile, non-spore-forming, yellow-pigmented and isolated from soil of a ginseng cultivation field in Pocheon T is allocated to the family Chitinophagaceae. Within the genus Province, South Korea. Gsoil 636 showed the ability to con- Flavisolibacter, the genomic DNA G + C content range is from vert Rb1 (one of the dominant active components of ginseng) to 42.7 to 46.4 mol%. Based on recent studies, this genus consists F2, and its whole genome was sequenced. The whole genome T of Flavisolibacter ginsenosidimutans Gsoil 636 consist of a of four species with validly published names and isolated from single circular chromosome of 5,079,621 bp, with 48.9% G + C various sources such as soil and water: Flavisolibacter gin- content. Of the 4,338 predicted genes, 4,251 were protein- sengiterrae, Flavisolibacter ginsengisoli (Yoon and Im, 2007), cording genes, 46 were RNAs, and 41 were pseudogenes. Using Flavisolibacter rigui (Baik et al., 2014), and Flavisolibacter T the complete genome sequence of the strain Gsoil 636 , we tropicus (Lee et al., 2016). identified several glycoside hydrolase-encoding genes that A ginsenoside transforming-positive, Gram-negative bacte- may be involved in the conversion of major ginsenosides into T rium, Flavisolibacter ginsenosidimutans Gsoil 636 , were minor ginsenosides and unexpectedly found antibiotic biosynthesis-encoding genes and excinuclease genes. isolated from soil of a ginseng cultivation field in Pocheon Province, South Korea. Based on the transform from major Keywords: Flavisolibacter ginsenosidimutans, complete genome, ginsenosides to minor ginsenosides (Zhao et al., 2015), F. ginseng soil, glycoside hydrolase, PacBio RS II T ginsenosidimutans Gsoil 636 was chosed for a whole genome study to identify the target functional genes. Whole genome The genus Flavisolibacter was first proposed by (Yoon and sequence analysis showed more than 70 glycoside hydrolases that could be involved in the conversion of ginsenosides. This *For correspondence. E-mail: [email protected]; strain is available from the host institute and from three culture Tel.: +82-31-670-5335; Fax: +82-31-670-5339 460 ∙ Keum et al.

T T T collections (= KCTC 22818 = JCM 18197 = KACC14277 ). with antibiotic biosynthesis, and multiple antibiotic resistance T Genomic DNA of F. ginsenosidimutans Gsoil 636 was proteins, excision endonuclease such as UvrABC subunits. extracted using a MagAttract HMW DNA kit (Qiagen) and was purified using the chloroform wash method (shared protocol; Nucleotide sequence accession number Pacific Biosciences). Genome sequencing was performed using The complete genome sequence of Flavisolibacter ginseno- a Pacific Biosciences RSII sequencing platform, with a 20 kb T sidimutans Gsoil 636 has been deposited in DDBJ/EMBL/ SMRTbellTM templates library, at DNA Link, Inc. Sequences NCBI GenBank under the accession number CP042433. were assembled using the HGAP3 (Pacific Biosciences) protocol and the sequencing depth was 194.66×. The genome sequence was annotated using the NCBI Prokaryotic Genome 적 요 Automatic Annotation Pipeline (http://www.ncbi.nlm.nih.gov/ books/NBK174280/) (DiCuccio et al., 2016). rRNAs and tRNAs 인삼 재배 토양으로부터 분리한 Flavisolibacter ginseno- T were predicted using rRNAmmer and tRNAscan-SE, respec- sidimutans Gsoil 636 균주의 유전체서열을 분석하였다. 균주 T tively. Gsoil 636 는 진세노사이드 Rb1을 F2로 전환하는 능력을 보 T T The complete genome of F. ginsenosidimutans Gsoil 636 여주었다. Gsoil 636 의 유전체는 G + C 비율이 48.9%이며, consisted of one circular chromosome of 5,079,621 bp, with 4,338개의 유전자와 4,251개의 단백질 코딩 유전자, 46개의 48.9% G + C content. Of the 4,338 predicted genes, 4,251 were RNA 유전자 그리고 41개의 위유전자를 포함한 단일 원형 염 protein-cording genes (CDSs), and 46 were RNA genes, and 41 색체로 구성되었으며 그 크기는 5,079,612 bp였다. 전체 DNA T pseudogenes. The majority of the protein coding genes (99.04%) 시컨싱을 통해 균주 Gsoil 636 이여러 타입의 글라이코시다 was assigned function, while the remaining predicted in Table 제 유전자를 가지고 있어 메이저 진세노사이드를 마이너진세 1. Analysis of the complete genome sequence showed many 노사이드로 전환하는 것을 확인하였다. 그 외에 항생제 생합 glycoside hydrolase-encoding genes, including 10 β-glucosidases, 성 유전자와 UV 관련 유전자도 가지고 있었다. 8 α-glucosidases, 23 β-xylosidases, 3 α-arabinofuranosidases, and 32 β-galactosidases, which may be related for its ability to convert ginsenosides. In addition, genome annotation revealed Acknowledgements other genes of interest, including nitrite reductase large subunits, vitamin B12-dependent ribonucleotide reductase, multiple This work was supported by grants from the National biosynthesis genes such as quinone biosynthesis C-methlylase Institute of Biological Resources, funded by the Ministry of

(UbiE), the cellulose synthase (CesA), and bacillithiol bio- Environment (No.NIBR201801106). synthesis deacetylase (BshB1), antibiotic related genes such as bleomycin resistance proteins, monooxygenase genes associated References T Table 1. General features of Flavisolibacter ginsenosidimutans Gsoil 636 Baik KS, Kim MS, Lee JH, Im WT, and Seong CN. 2014. Flavi- Features Chromosome solibacter rigui sp. nov., isolated from freshwater of an artificial Genome size (bp) 5,079,621 reservoir and emended description of the genus Flavisolibacter. DNA coding region (bp) 4,977,748 Int. J. Syst. Evol. Microbiol. 64, 4038–4042. G + C content (%) 48.9 DiCuccio M, Zaslavsky L, Chetvernin V, Ostell J, Badretdin A, Total genes 4,338 Tatusova T, Lomsadze A, Borodovsky M, Nawrocki EP, and Pruitt KD. 2016. NCBI prokaryotic genome annotation pipeline. Pseudo genes 41 Nucleic Acids Res. 44, 6614–6624. Protein-cording genes 4,251 Lee JJ, Kang MS, Kim GS, Lee CS, Lim SY, Lee JD, Roh SH, Kang Number of rRNA genes (5S, 16S, 23S) 3 (1, 1, 1) HR, Ha JM, Bae SJ, et al. 2016. Flavisolibacter tropicus sp. Number of tRNA genes 40 nov., isolated from tropical soil. Int. J. Syst. Evol. Microbiol. 66, Number of ncRNA genes 3 3413–3419.

미생물학회지 제55권 제4호 T Complete genome sequence of F. ginsenosidimutans Gsoil 636 ∙ 461

Yoon MH and Im WT. 2007. Flavisolibacter ginsengiterrae gen. nov., Zhao Y, Liu Q, Kang MS, Jin FX, Yu HS, and Im WT. 2015. sp. nov. and Flavisolibacter ginsengisoli sp. nov., isolated from Flavisolibacter ginsenosidimutans sp. nov., with ginsenoside- ginseng cultivating soil. Int. J. Syst. Evol. Microbiol. 57, 1834– converting activity isolated from soil used for cultivating 1839. ginseng. Int. J. Syst. Evol. Microbiol. 65, 4868–4872.

Korean Journal of Microbiology, Vol. 55, No. 4