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Journal of Bacteriology, January 2003, p. 601-609, Vol. 185, No. 2
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.2.601-609.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Enhanced Expression of S-Adenosylmethionine Synthetase Causes Overproduction of Actinorhodin in Streptomyces coelicolor A3(2){dagger}

Susumu Okamoto, Alexander Lezhava, Takeshi Hosaka, Yoshiko Okamoto-Hosoya, and Kozo Ochi*

National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan

Received 26 July 2002/ Accepted 9 October 2002

We found that a 46-kDa protein is highly expressed in an actinorhodin-overproducing Streptomyces coelicolor A3(2) mutant (KO-179), which exhibited a low-level resistance to streptomycin. The protein was identified as S-adenosylmethionine (SAM) synthetase, which is a product of the metK gene. Enzyme assay revealed that SAM synthetase activity in strain KO-179 was 5- to 10-fold higher than in wild-type cells. The elevation of SAM synthetase activity was found to be associated with an increase in the level of intracellular SAM. RNase protection assay revealed that the metK gene was transcribed from two distinct promoters (p1 and p2) and that enhanced expression of the MetK protein in the mutant strain KO-179 was attributed to elevated transcription from metKp2. Strikingly, the introduction of a high-copy-number plasmid containing the metK gene into wild-type cells resulted in a precocious hyperproduction of actinorhodin. Furthermore, the addition of SAM to the culture medium induced Act biosynthesis in wild-type cells. Overexpression of metK stimulated the expression of the pathway-specific regulatory gene actII-ORF4, as demonstrated by the RNase protection assay. The addition of SAM also caused hyperproduction of streptomycin in Streptomyces griseus. These findings implicate the significant involvement of intracellular SAM in initiating the onset of secondary metabolism in Streptomyces.

* Corresponding author. Mailing address: National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan. Phone: 81-298-38-8125. Fax: 81-298-38-7996. E-mail: kochi{at}affrc.go.jp.

{dagger} This paper is dedicated to David A. Hopwood upon his retirement from the John Innes Institute.

Journal of Bacteriology, January 2003, p. 601-609, Vol. 185, No. 2
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.2.601-609.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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