Protein & Cell

, Volume 4, Issue 11, pp 854–862

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

Research Article Protein & Cell

Abstract

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.

Keywords

Clostridium acetobutylicum mutation rates hypermutable cells artificial control 

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Copyright information

© Higher Education Press and Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  1. 1.CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of MicrobiologyChinese Academy of SciencesBeijingChina
  2. 2.University of Chinese Academy of SciencesBeijingChina

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