Role of the Cro repressor carboxy terminal domain and flexible dimer linkage in operator and nonspecific DNA binding

• Published in: Biochemistry (Easton) Vol. 29; no. 39; pp. 9241 - 9249
• Main Authors: Hubbard, Adrian J, Bracco, Laurent P, Eisenbeis, Scott J, Gayle, Richard B, Beaton, Graham, Caruthers, Marvin H
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 02.10.1990
• Subjects: Amino Acid Sequence; Base Sequence; Biological and medical sciences; Cloning, Molecular; DNA - metabolism; Fundamental and applied biological sciences. Psychology; Genetics; Microbiology; Molecular Sequence Data; Mutation; Operator Regions, Genetic; phage lambda; Repressor Proteins - genetics; Virology; Site directed mutagenesis; Association constant; Molecular interaction; Transcription repressor; Virus; C terminal-Sequence; Lambda phage group; Transcription operator; Styloviridae; DNA; Molecular complex; Phage lambda; Dimerization; Phage; Hydrogen bond
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi00491a019

A series of mutations comprising single and multiple substitutions, deletions, and extensions within the carboxy-terminal domain of the bacteriophage lambda Cro repressor have been constructed. These mutations generally affect the affinity of repressor for specific and nonspecific DNA. Additionally, substitution of the carboxy-terminal alanine with several amino acids capable of hydrogen-bonding interactions leads to improved specific binding affinities. A mutation is also described whereby cysteine links the two Cro monomers by a disulfide bond. As a consequence, a significant improvement in nonspecific binding and a concomitant reduction in specific binding are observed with this mutant. These results provide evidence that the carboxy terminus of Cro repressor is an important DNA binding domain and that a flexible connection between the two repressor monomers is a critical factor in modulating the affinity of wild-type repressor for DNA.