PMID:11124943
| Citation |
Loh, T, Murphy, KC and Marinus, MG (2001) Mutational analysis of the MutH protein from Escherichia coli. J. Biol. Chem. 276:12113-9 |
|---|---|
| Abstract |
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). |
| Links |
PubMed Online version:10.1074/jbc.M007935200 |
| Keywords |
Base Sequence; DNA Primers; DNA Repair Enzymes; DNA-Binding Proteins/chemistry; DNA-Binding Proteins/genetics; DNA-Binding Proteins/metabolism; Endodeoxyribonucleases/chemistry; Endodeoxyribonucleases/genetics; Endodeoxyribonucleases/metabolism; Escherichia coli/metabolism; Escherichia coli Proteins; Models, Molecular; Mutagenesis, Site-Directed; Protein Conformation |
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