Identification of specific amino acid residues in the E. coli β processivity clamp involved in interactions with DNA polymerase III, UmuD and UmuD

• Published in: DNA repair 2004-03, Vol.3 (3), p.301-312
• Main Authors: Duzen, Jill M., Walker, Graham C., Sutton, Mark D.
• Format: Article
• Language: English
• Subjects: Amino Acids - metabolism; Bacterial Proteins - metabolism; Bacteriology; Biological and medical sciences; Blotting, Western; Chromatography, Affinity; Cross-Linking Reagents - metabolism; Densitometry; DNA Polymerase III - metabolism; DNA polymerase management; DNA Primers; DNA Repair - physiology; DNA replication; DNA-Directed DNA Polymerase; dnaN; Escherichia coli; Escherichia coli - genetics; Escherichia coli - physiology; Escherichia coli Proteins - metabolism; Formaldehyde; Fundamental and applied biological sciences. Psychology; Glutaral; Growth, nutrition, cell differenciation; Microbiology; Molecular and cellular biology; Molecular genetics; Mutagenesis, Site-Directed - physiology; Mutagenesis. Repair; Protein Binding; Translesion DNA synthesis; umuDC; β sliding clamp
• ISSN: 1568-7864; 1568-7856
• DOI: 10.1016/j.dnarep.2003.11.008

Variants of a pentapeptide sequence (QL[S/F]LF), referred to as the eubacterial clamp-binding motif, appear to be required for certain proteins to bind specifically to the Escherichia coli β sliding clamp, apparently by making contact with a hydrophobic pocket located at the base of the C-terminal tail of each β protomer. Although both UmuC (DNA pol V) and the α catalytic subunit of DNA polymerase III (pol III) each bear a reasonable match to this motif, which appears to be required for their respective interactions with the clamp, neither UmuD not UmuD′ do. As part of an ongoing effort to understand how interactions involving the different E. coli umuDC gene products and components of DNA polymerase III help to coordinate DNA replication with a DNA damage checkpoint control and translesion DNA synthesis (TLS) following DNA damage, we characterized the surfaces on β important for its interactions with the two forms of the umuD gene product. We also characterized the surface of β important for its interaction with the α catalytic subunit of pol III. Our results indicate that although UmuD, UmuD′ and α share some common contacts with β, each also makes unique contacts with the clamp. These findings suggest that differential interactions of UmuD and UmuD′ with β impose a DNA damage-responsive conditionality on how β interacts with the translesion DNA polymerase UmuC. This is formally analogous to how post-translational modification of the eukaryotic PCNA clamp influences mutagenesis. We discuss the implications of our findings in terms of how E. coli might coordinate the actions of the umuDC gene products with those of pol III, as well as for how organisms in general might manage the actions of their multiple DNA polymerases.