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Journal of Bacteriology, October 2000, p. 5807-5812, Vol. 182, No. 20
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Characterization of Bacteriophage Lambda Excisionase Mutants Defective in DNA Binding

Eun Hee Cho,1 Renato Alcaraz Jr.,2 Richard I. Gumport,3 and Jeffrey F. Gardner2,*

Department of Science Education, Chosun University, Kwangju, Korea,1 and Department of Microbiology2 and Department of Biochemistry and College of Medicine,3 University of Illinois, Urbana, Illinois

Received 22 May 2000/Accepted 28 July 2000

The bacteriophage lambda  excisionase (Xis) is a sequence-specific DNA binding protein required for excisive recombination. Xis binds cooperatively to two DNA sites arranged as direct repeats on the phage DNA. Efficient excision is achieved through a cooperative interaction between Xis and the host-encoded factor for inversion stimulation as well as a cooperative interaction between Xis and integrase. The secondary structure of the Xis protein was predicted to contain a typical amphipathic helix that spans residues 18 to 28. Several mutants, defective in promoting excision in vivo, were isolated with mutations at positions encoding polar amino acids in the putative helix (T. E. Numrych, R. I. Gumport, and J. F. Gardner, EMBO J. 11:3797-3806, 1992). We substituted alanines for the polar amino acids in this region. Mutant proteins with substitutions for polar amino acids in the amino-terminal region of the putative helix exhibited decreased excision in vivo and were defective in DNA binding. In addition, an alanine substitution at glutamic acid 40 also resulted in altered DNA binding. This indicates that the hydrophilic face of the alpha -helix and the region containing glutamic acid 40 may form the DNA binding surfaces of the Xis protein.

* Corresponding author. Mailing address: B103 Chemical and Life Science Lab., 601 S. Goodwin Ave., Urbana, IL 61801. Phone: (217) 333-7287. Fax: (217) 244-6697. E-mail: jeffgard{at}uiuc.edu.

Journal of Bacteriology, October 2000, p. 5807-5812, Vol. 182, No. 20
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

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