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Phase variation and the Hin protein: in vivo activity measurements, protein overproduction, and purification

Bruist, Michael F. and Simon, Melvin I. (1984) Phase variation and the Hin protein: in vivo activity measurements, protein overproduction, and purification. Journal of Bacteriology, 159 (1). pp. 71-79. ISSN 0021-9193.

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The alternate expression of the Salmonella flagellin genes H1 and H2 is controlled by the orientation of a 995-base-pair invertible segment of DNA located at the 5' end of the H2 gene. The hin gene, which is encoded within the invertible region, is essential for the inversion of this DNA segment. We cloned the hin gene into Escherichia coli and placed it under the control of the PL promoter of bacteriophage lambda. These cells overproduced the Hin protein. In vivo inversion activity was measured by using a recombinant lambda phage which contains the H2 and lacZ genes under the control of the invertible region. Using this phage, we showed that the amount of inversion activity is proportional to the amount of Hin protein in the cell. An inactive form of the protein was purified by using the unusual solubility properties of the overproduced protein. The amino acid composition of the protein agreed with the DNA sequence of the hin gene. Antibodies were made to the isolated protein. These antibodies cross-reacted with two other unidentified E. coli proteins.

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Additional Information:Copyright © 1984 by the American Society for Microbiology. Received 10 February 1984/Accepted 9 April 1984 We thank P. Price for performing the amino acid analysis of the Hin protein. We also thank M. Silverman, G. Mandel, J. Zieg, and A. Hoyt for providing us with phage, strains, and plasmids. Special thanks to E. Phizicky for helpful discussions and critical evaluation of the manuscript. This work was supported by National Science Foundation grant PCM 8209295.
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Deposited On:31 Mar 2006
Last Modified:02 Oct 2019 22:52

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