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Journal of Bacteriology, January 2004, p. 1-7, Vol. 186, No. 1
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.1.1-7.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

The Preferred Substrate for RecA-Mediated Cleavage of Bacteriophage 434 Repressor Is the DNA-Bound Dimer

David R. Pawlowski and Gerald B. Koudelka*

Department of Biological Sciences, University at Buffalo, Buffalo, New York 14260

Received 11 June 2003/ Accepted 1 October 2003

Induction of a lysogen of a lambdoid bacteriophage usually involves RecA-stimulated autoproteolysis of the bacteriophage repressor protein. Previous work on the phage repressors showed that the monomeric form of the protein is the target of RecA. Our previous work indicated that in the case of bacteriophage 434, virtually none of the repressor is present as a monomer in vivo. Hence, if the repressor in a lysogen is present as a dimer, how can RecA-stimulated autoproteolysis play a role in bacteriophage induction? We examined this question by determining the rate of RecA-stimulated 434 repressor cleavage as a function of repressor concentration and added DNA. Our results show that binding of 434 repressor to a specific DNA binding site dramatically increases the velocity of repressor autocleavage compared to the velocity of cleavage of the monomer and concentration-induced dimer. DNA binding-deficient hemidimers formed between the intact repressor and its C-terminal domain fragment have a lower rate of cleavage than DNA-bound dimers. These results show that the DNA-bound 434 repressor dimer, which is the form of the repressor that is required for its transcriptional regulatory functions, is the preferred form for RecA-stimulated autocleavage. We also show that the rate of repressor autocleavage is influenced by the sequence of the bound DNA. Kinetic analysis of the autocleavage reaction indicated that the DNA sequence influences the velocity of 434 repressor autocleavage by affecting the affinity of the repressor-DNA complex for RecA, not the chemical cleavage step. Regardless of the mechanism, the finding that the presence and precise sequence of DNA modulate the autocleavage reaction shows that DNA allosterically affects the function of 434 repressor.

* Corresponding author. Mailing address: Department of Biological Sciences, University at Buffalo, Cooke Hall, North Campus, Buffalo, NY 14260-1300. Phone: (716) 645-2363, ext. 158. Fax: (716) 645-2975. E-mail: koudelka{at}acsu.buffalo.edu.

Journal of Bacteriology, January 2004, p. 1-7, Vol. 186, No. 1
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.1.1-7.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • McCabe, B. C., Pawlowski, D. R., Koudelka, G. B. (2005). The Bacteriophage 434 Repressor Dimer Preferentially Undergoes Autoproteolysis by an Intramolecular Mechanism. J. Bacteriol. 187: 5624-5630 [Abstract] [Full Text]  


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