Mutational analysis of the MutH protein from Escherichia coli
Graduate School of Biomedical Sciences; Department of Molecular Genetics and Microbiology; Department of Biochemistry and Molecular Pharmacology
Life Sciences | Medicine and Health Sciences
Site-directed mutagenesis was performed on several areas of MutH based on the similarity of MutH and PvuII structural models. The aims were to identify DNA-binding residues; to determine whether MutH has the same mechanism for DNA binding and catalysis as PvuII; and to localize the residues responsible for MutH stimulation by MutL. No DNA-binding residues were identified in the two flexible loop regions of MutH, although similar loops in PvuII are involved in DNA binding. Two histidines in MutH are in a similar position as two histidines (His-84 and His-85) in PvuII that signal for DNA binding and catalysis. These MutH histidines (His-112 and His-115) were changed to alanines, but the mutant proteins had wild-type activity both in vivo and in vitro. The results indicate that the MutH signal for DNA binding and catalysis remains unknown. Instead, a lysine residue (Lys-48) was found in the first flexible loop that functions in catalysis together with the three presumed catalytic amino acids (Asp-70, Glu-77, and Lys-79). Two deletion mutations (MutHDelta224 and MutHDelta214) in the C-terminal end of the protein, localized the MutL stimulation region to five amino acids (Ala-220, Leu-221, Leu-222, Ala-223, and Arg-224).
DOI of Published Version
J Biol Chem. 2001 Apr 13;276(15):12113-9. Epub 2000 Dec 21. Link to article on publisher's site
The Journal of biological chemistry
Loh T, Murphy KC, Marinus MG. (2001). Mutational analysis of the MutH protein from Escherichia coli. Morningside Graduate School of Biomedical Sciences Student Publications. https://doi.org/10.1074/jbc.M007935200. Retrieved from https://escholarship.umassmed.edu/gsbs_sp/779