Functional interaction between MutL and 3′–5′ Exonuclease X in Escherichia coli

• Published in: Archives of biochemistry and biophysics 2010-10, Vol.502 (1), p.39-43
• Main Authors: Cheng, Fang, Hou, Jian, Chen, Yuan-Yuan, Zhou, Ying, Zhang, Hong-Tai, Bi, Li-Jun, Zhang, Xian-En
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphatases - chemistry; Adenosine Triphosphatases - genetics; Adenosine Triphosphatases - metabolism; Adenosine Triphosphate - metabolism; Amino Acid Sequence; Base Sequence; Blotting, Far-Western; DNA Breaks, Double-Stranded; DNA Mismatch Repair; DNA, Bacterial - genetics; DNA, Bacterial - metabolism; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Exonucleases - chemistry; Exonucleases - genetics; Exonucleases - metabolism; ExoX; Ionic interaction; Models, Biological; Mutant Proteins - genetics; Mutant Proteins - metabolism; MutL; MutL Proteins; Plasmids - genetics; Promotion mechanism; Protein Interaction Mapping; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Surface Plasmon Resonance
• ISSN: 0003-9861; 1096-0384
• DOI: 10.1016/j.abb.2010.07.011

► MutL physically interacts with Exonuclease X. ► MutL stimulates the exonuclease activity of ExoX. ► Stimulation of ExoX by MutL is ATP-independent. ► Stimulation of ExoX does not require the DNA-binding activity of MutL. ► The 335 N-terminal amino acids of MutL are essential for functional interactions. Exonuclease X is a 3′–5′ distributive exonuclease that functions in DNA recombination and repair. It undergoes multiple rounds of binding, hydrolysis, and release to degrade long substrate molecules and thus is very inefficient. In order to identify a cofactor that elevates the excision activity of ExoX, we screened many proteins involved in repair and recombination. We observed that MutL greatly promoted the exonuclease activity of ExoX, and then verified the interaction between MutL and ExoX using SPR and Far-Western analysis. This promotion is independent of ATP and the DNA-binding activity of MutL. We constructed two deletion mutants to analyze this interaction and its regulation of ExoX activity, and found that this functional interaction with ExoX is mainly due to ionic interactions with the N-terminus of MutL. This adds a new role to MutL and gives a clue to MutL’s possible regulation on other DnaQ family exonuclease members.