Abstract
We report that Corynebacterium glutamicum colonies exhibit a developmental transition in culture. When cultured on a routinely used complete medium (CM2B), this bacterium first formed a flat translucent colony. Subsequently, some parts of this colony expanded to form small spherical yellow colonies that finally developed into a single large yellow colony. The small flat colony consisted of long thick cells, which were occasionally V or Y shaped, while the large yellow colony consisted of short small rods. A similar colony development pattern was observed in Corynebacterium ammoniagenes and Corynebacterium callunae. Analysis following shotgun cloning revealed that the introduction of a multi-copy-number plasmid carrying amtR, a global transcriptional regulator for nitrogen metabolism, into C. glutamicum cells induced precocious colony development. An amtR-null C. glutamicum mutant exhibited delayed development. Detailed observations of C. glutamicum cells cultured on CM2B medium containing buffers at various pH values revealed that the colony growth was rapid at a pH value of 6.4 or higher and slow but distinct at a pH of less than 6.4. This pH threshold increased to 6.8 following the addition of 0.1% glucose into the medium.
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Acknowledgments
We thank Hisashi Kawasaki for providing useful information and Shoichi Amano for his assistance with regard to the scanning electron microscopic observations. This study was supported by the High-Tech Research Centre Project of the Ministry of Education, Culture, Sports, Science and Technology, Japan.
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Supplemental Figure S1
A representative result of proteomic analysis. Total proteins extracted from C. glutamicum wild-type cells comprising small translucent (ST) and large yellow (LY) colonies were analyzed and compared. Cells were grown for 5 days on CM2G (for former colonies) and CM2B (for latter colonies), washed with 50 mM Tris–HCl (pH 7.0) buffer and disrupted by sonication. Supernatant obtained after centrifuging sonicated cell suspension at 12,000×g for 10 min (containing 1.0 mg protein) was further treated with a 2-D Clean-Up Kit and subjected to 2-D gel electrophoresis using Multiphore II system. The method of electrophoresis followed the manufacturer’s standard protocol. Proteins were stained with Flamingo fluorescent dye and visualized by using a Typhoon image analyzer. All systems and materials used were products of GE Healthcare (PDF 1.95 MB).
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Takano, H., Shimizu, A., Shibosawa, R. et al. Characterization of developmental colony formation in Corynebacterium glutamicum . Appl Microbiol Biotechnol 81, 127–134 (2008). https://doi.org/10.1007/s00253-008-1622-z
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DOI: https://doi.org/10.1007/s00253-008-1622-z