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Applied and Environmental Microbiology, November 2006, p. 7074-7082, Vol. 72, No. 11
0099-2240/06/$08.00+0     doi:10.1128/AEM.01334-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Analysis of Novel Soluble Chromate and Uranyl Reductases and Generation of an Improved Enzyme by Directed Evolution

Y. Barak,1 D. F. Ackerley,1,{dagger} C. J. Dodge,2 L. Banwari,1,{ddagger} C. Alex,1 A. J. Francis,2 and A. Matin1*

Department of Microbiology and Immunology, Sherman Fairchild Science Building, Stanford University School of Medicine, 299 Campus Drive, Stanford, California 94305,1 Environmental Sciences Department, Brookhaven National Laboratory, Upton, New York2

Received 9 June 2006/ Accepted 27 July 2006

Most polluted sites contain mixed waste. This is especially true of the U.S. Department of Energy (DOE) waste sites which hold a complex mixture of heavy metals, radionuclides, and organic solvents. In such environments enzymes that can remediate multiple pollutants are advantageous. We report here evolution of an enzyme, ChrR6 (formerly referred to as Y6), which shows a markedly enhanced capacity for remediating two of the most serious and prevalent DOE contaminants, chromate and uranyl. ChrR6 is a soluble enzyme and reduces chromate and uranyl intracellularly. Thus, the reduced product is at least partially sequestered and nucleated, minimizing the chances of reoxidation. Only one amino acid change, Tyr128Asn, was responsible for the observed improvement. We show here that ChrR6 makes Pseudomonas putida and Escherichia coli more efficient agents for bioremediation if the cellular permeability barrier to the metals is decreased.


* Corresponding author. Mailing address: Department of Microbiology and Immunology, 299 Campus Drive West, Sherman Fairchild D317, Stanford University, Stanford, CA 94305-5124. Phone: (650) 725-4745. Fax: (650) 725-6757. E-mail: a.matin{at}stanford.edu.

{dagger} Present address: School of Biological Sciences, Victoria University, Kelburn Parade, Wellington, New Zealand.

{ddagger} Present address: The Energy and Resources Institute, Darbari Seth Block, IHC Complex, Lodhi Road, New Delhi 110 003, India.


Applied and Environmental Microbiology, November 2006, p. 7074-7082, Vol. 72, No. 11
0099-2240/06/$08.00+0     doi:10.1128/AEM.01334-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.




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