Volume 4 Issue 11
Nov.  2013
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Guodong Luan, Zhen Cai, Fuyu Gong, Hongjun Dong, Zhao Lin, Yanping Zhang, Yin Li. Developing controllable hypermutable Clostridium cells through manipulating its methyl-directed mismatch repair system[J]. Protein&Cell, 2013, 4(11): 854-862. doi: 10.1007/s13238-013-3079-9
Citation: Guodong Luan, Zhen Cai, Fuyu Gong, Hongjun Dong, Zhao Lin, Yanping Zhang, Yin Li. Developing controllable hypermutable Clostridium cells through manipulating its methyl-directed mismatch repair system[J]. Protein&Cell, 2013, 4(11): 854-862. doi: 10.1007/s13238-013-3079-9

Developing controllable hypermutable Clostridium cells through manipulating its methyl-directed mismatch repair system

doi: 10.1007/s13238-013-3079-9
Funds:

This research was supported by grants from the National Natural Science Foundation of China (Grant No. 31270107), the National Basic Research Program of China (973 Program) (No. 2011CBA00800) and the National High Technology Research and Development Program (863 Program) (No. 2011AA02A208). Yanping Zhang is supported by grants from the Knowledge Innovation Program of the Chinese Academy of Sciences (No. KSCX2-EW-Q-14).

  • Received Date: 2013-09-05
  • Rev Recd Date: 2013-09-25
  • Development of controllable hypermutable cells can greatly benefit understanding and harnessing microbial evolution. However, there have not been any similar systems developed for Clostridium, an important bacterial genus. Here we report a novel two-step strategy for developing controllable hypermutable cells of Clostridium acetobutylicum, an important and representative industrial strain. Firstly, the mutS/L operon essential for methyldirected mismatch repair (MMR) activity was inactivated from the genome of C. acetobutylicum to generate hypermutable cells with over 250-fold increased mutation rates. Secondly, a proofreading control system carrying an inducibly expressed mutS/L operon was constructed. The hypermutable cells and the proofreading control system were integrated to form a controllable hypermutable system SMBMutC, of which the mutation rates can be regulated by the concentration of anhydrotetracycline (aTc). Duplication of the miniPthl-tetR module of the proofreading control system further significantly expanded the regulatory space of the mutation rates, demonstrating hypermutable Clostridium cells with controllable mutation rates are generated. The developed C. acetobutylicum strain SMBMutC2 showed higher survival capacities than the control strain facing butanol-stress, indicating greatly increased evolvability and adaptability of the controllable hypermutable cells under environmental challenges.
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