Basic Research
Copyright ©The Author(s) 2005. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Oct 21, 2005; 11(39): 6152-6158
Published online Oct 21, 2005. doi: 10.3748/wjg.v11.i39.6152
Identification of anrF gene, a homology of admM of andrimid biosynthetic gene cluster related to the antagonistic activity of Enterobacter cloacae B8
Xu-Ping Yu, Jun-Li Zhu, Xue-Ping Yao, Shi-Cheng He, Hai-Ning Huang, Wei-Liang Chen, Yong-Hao Hu, De-Bao Li
Xu-Ping Yu, Jun-Li Zhu, Xue-Ping Yao, Shi-Cheng He, Veterinary Medicine Department, Animal Science College, Zhejiang University, Hangzhou 310029, Zhejiang Province, China
Xu-Ping Yu, Hai-Ning Huang, Wei-Liang Chen, De-Bao Li, Institute of Biotechnology, Zhejiang University, Hangzhou 310029, Zhejiang Province, China
Jun-Li Zhu, College of Food Science, Biotechnology and Environmental Engineering, Zhejiang Gongshang University, Hangzhou 310035, Zhejiang Province, China
Xue-Ping Yao, Yong-Hao Hu, Veterinary Medicine Department, Animal Science College, Gansu Agricultural University, Lanzhou 730070, Gansu Province, China
Author contributions: All authors contributed equally to the work.
Supported by the National Natural Science Foundation of China, No. 39870034; the National High Technology Research and Development Program of China, the 863 Program, No. 104-04-01-01; and the Major Project of Science and Technology Development of Zhejiang Province, No. 021102529
Correspondence to: Dr. Xu-Ping Yu, Veterinary Medicine Department, Animal Science College, Zhejiang University, Hangzhou 310029, Zhejiang Province, China. xpyu@zju.edu.cn
Telephone: +86-571-86971096 Fax: +86-571-86971096
Received: January 24, 2005
Revised: April 8, 2005
Accepted: April 11, 2005
Published online: October 21, 2005
Abstract

AIM: To identify the gene (s) related to the antagonistic activity of Enterobacter cloacae B8 and to elucidate its antagonistic mechanism.

METHODS: Transposon-mediated mutagenesis and tagging method and cassette PCR-based chromosomal walking method were adopted to isolate the mutant strain(s) of B8 that lost the antagonistic activity and to clone DNA fragments around Tn5 insertion site. Sequence compiling and open reading frame (ORF) finding were done with DNAStar program and homologous sequence and conserved domain searches were performed with BlastN or BlastP programs at www.ncbi.nlm.nih.gov. To verify the gene involved in the antagonistic activity, complementation of a full-length clone of the anrF gene to the mutant B8F strain was used.

RESULTS: A 3 321 bp contig around the Tn5 insertion site was obtained and an ORF of 2 634 bp in length designated as anrFgene encoding for a 877 aa polyketide synthase-like protein was identified. It had a homology of 83% at the nucleotide level and 79% ID/87% SIM at the protein level, to the admM gene of Pantoea agglomerans andrimid biosynthetic gene cluster (AY192157). The Tn5 was inserted at 2 420 bp of the gene corresponding to the COG3319 (the thioesterase domain of type I polyketide synthase) coding region on B8F. The antagonistic activity against Xanthomonas oryzae pv. oryzae was resumed with complementation of the full-length anrF gene to the mutant B8F.

CONCLUSION: The anrF gene obtained is related to the antagonistic activity of B8, and the antagonistic substances produced by B8 are andrimid and/or its analogs.

Keywords: Enterobacter cloacae B8; Antagonistic mechanism; anrF gene; Andrimid biosynthetic gene cluster