Rapid functional screening of Streptomyces coelicolor regulators by use of a pH indicator and application to the MarR-like regulator AbsC.
To elucidate the function of an unknown regulator in Streptomyces, differences in phenotype and antibiotic production between a deletion mutant and a wild-type strain (WT) were compared. These differences are easily hidden by complex media. To determine the specific nutrient conditions that reveal such differences, we used a multiwell method containing different nutrients along with bromothymol blue. We found several nutrients that provide key information on characterization conditions. By comparing the growth of wild-type and mutant strains on screened nutrients, we were able to measure growth, organic acid production, and antibiotic production for the elucidation of regulator function. As a result of this method, a member of the MarR-like regulator family, SCO5405 (AbsC), was newly characterized to control pyruvate dehydrogenase in Streptomyces coelicolor. Deletion of SCO5405 increased the pH of the culture broth due to decreased production of organic acids such as pyruvate and alpha-ketoglutarate and increased extracellular actinorhodin (ACT) production in minimal medium containing glucose and alanine (MMGA). This method could therefore be a high-throughput method for the characterization of unknown regulators.
Yang
Yung-Hun
YH
School of Chemical and Biological Engineering, Institute of Bioengineering, Seoul National University, Seoul, Republic of Korea.
Song
Eunjung
E
Lee
Bo-Rahm
BR
Kim
Eun-jung
EJ
Park
Sung-Hee
SH
Kim
Yun-Gon
YG
Lee
Chang-Soo
CS
Kim
Byung-Gee
BG
D000900:Anti-Bacterial Agents / Q000096:biosynthesis; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001431:Bacteriological Techniques / Q000379:methods*; D002264:Carboxylic Acids / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry*; D017353:Gene Deletion; D057166:High-Throughput Screening Assays; D006863:Hydrogen-Ion Concentration; D048372:Streptomyces coelicolor / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002264:Carboxylic Acids; D003470:Culture Media; D014157:Transcription Factors
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