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Applied and Environmental Microbiology, July 2007, p. 4515-4521, Vol. 73, No. 14
0099-2240/07/$08.00+0     doi:10.1128/AEM.02857-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Novel Redox Potential-Based Screening Strategy for Rapid Isolation of Klebsiella pneumoniae Mutants with Enhanced 1,3-Propanediol-Producing Capability ^,

Chenyu Du,^1, Yanping Zhang,^1,2 Yin Li,² and Zhu'an Cao^1*

Institute of Biochemical Engineering, Department of Chemical Engineering, Tsinghua University, Beijing 100084, People's Republic of China,¹ Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, People's Republic of China²

Received 8 December 2006/ Accepted 12 May 2007

This report describes a novel redox potential (oxidoreduction potential [ORP])-based screening strategy for the isolation of mutants of Klebsiella pneumoniae which have an increased ability to produce 1,3-propanediol (1,3-PD). This method can be described as follows: first, to determine an ORP range which is preferred for the wild-type strain to grow and to produce 1,3-PD; second, to subject a chemically mutagenized culture to a reduced ORP level which is deleterious for the wild-type strain. Colonies that showed high specific growth rates at deleterious ORP levels were selected, and their abilities to produce 1,3-PD were investigated. In an ORP-based screening experiment where the ORP was controlled at –280 mV, 4 out of 11 isolated strains were recognized as positive mutant strains. A mutant which is capable of producing higher concentrations of 1,3-PD was subjected to fed-batch fermentations for further characterization. Its preferred ORP level (–280 mV) was significantly lower than that of its parent (–190 mV). The highest 1,3-PD concentration of the mutant was 915 mmol liter^–1, which was 63.1% higher than that of the parent. Metabolic-flux analysis suggested that the intracellular reductive branch of the mutant was strengthened, which improved 1,3-PD biosynthesis. The procedure and results presented here provide a novel method of screening for strains with improved product formation.

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* Corresponding author. Mailing address: Department of Chemical Engineering, Tsinghua University, Beijing 100084, People's Republic of China. Phone: 86-10-62785514. Fax: 86-10-62770304. E-mail: cza-dce{at}mail.tsinghua.edu.cn

Published ahead of print on 18 May 2007.

Supplemental material for this article may be found at http://aem.asm.org/.

Present address: Satake Centre for Grain Process Engineering, School of Chemical Engineering and Analytical Science, The University of Manchester, Manchester M60 1QD, United Kingdom.

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Applied and Environmental Microbiology, July 2007, p. 4515-4521, Vol. 73, No. 14
0099-2240/07/$08.00+0     doi:10.1128/AEM.02857-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

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