kinase structurally similar to YycG, by docking analysis of aranorosinol B against EnvZ using Autodock v3.0.S. These results demonstrated that aranorosinol B bound to ATP-binding region of YycG and inhibited autophosphorylation of YycG (Fig. 2). The ts growth assay described here is presently being used in our laboratory for high-throughput screening for the identification of HK inhibitors and is of value in the subsequent stages of lead identification involving iterative rounds of chemical refinement.
ACKNOWLEDGMENTS
I wish to express my sincerest gratitude to Professor Dr. Ryutaro Utsumi, Department of Bioscience and Biotechnolgy, Graduate School of Agriculture, Kinki University, for kind guidance, valuable suggestion and discussions, and continuous encouragement throughout the course of this work and critical reading of the manuscript.
Special thanks are due to Drs. Tadashi Okamoto and Tamo Fukamizo, Professor of Department of Bioscience and Biotechnolgy, Graduate School of Agriculture, Kinki University, for reading the entire text in its original form.
I also wish to express my sincerest acknowlegment to Assosiate Professor Dr.
Hiroyuki Tanabe, Department of Agicultural Chemistry, Kinki University, and Assistant Professor Ph. D. Kaneyoshi Yamamoto, Department of Agicultural Chemistry, Kinki University, for their useful discussions and technical advices.
I am grateful to Daniel Ladant for generously supplying me with derivatives of pT18 and pT25 plasmids and DHPI strains, and Robert Novy for generous supplying me with pKWY2428 plasmid.
I would also like to thank Dr. Hirofumi Yoshikawa, Laboratory of Microbiology and Molecula Genetics, Tokyo University of Agricultute, for providing B. subtilis 168; Dr.
Mitsuhiko Itaya, Mitsubishi Kasei Institute of Life Sciences, for providing drug-resistant genes for B. subtilis, pBEST501 and pBEST4C.
I thank Drs. Motohiro Hino, Akihiko Fujie, Yasuhisa Tsurumi and Shigehiro Takase, Exploratory Research Laboratries Fujisawa Pharmaceutical Co., Ltd., for supplying acetone extracts from 4000 microbe, and identification of G. dankaliensis and aranorosinol B.
Special thanks are due to Drs. Masayuki Matsushita, Assistant Professor of The Scripps Research Institute, for the docking experiments of aranorosinol B against EnvZ and valuable suggestion and technical advices.
Finally, I am great indebted to Yoshiki Hashimoto, Yoshihito Umemoto, Daisuke Tatebe, and other members of Department of Bioscience and Biotechnolgy, Graduate School of Agriculture and Department of Agicultural Chemistry, Kinki University.
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