hMutSalpha forms an ATP-dependent complex with hMutLalpha and hMutLbeta on DNA.
The DNA binding properties of hMutSalpha and hMutLalpha and complex formation of hMutSalpha with hMutLalpha and hMutLbeta were investigated using binding experiments on magnetic bead-coupled DNA substrates with nuclear extracts as well as purified proteins. hMutSalpha binding to homoduplex DNA was disrupted by lower NaCl concentrations than hMutSalpha binding to a mismatch. ATP markedly reduced the salt resistance of hMutSalpha binding but hMutSalpha still retained affinity for heteroduplexes. hMutSalpha formed a complex with hMutLalpha and hMutLbeta on DNA in the presence of ATP. This complex only formed on 81mer and not 32mer DNA substrates. Complex formation was enhanced by a mismatch in the DNA substrate, and hMutLalpha and hMutLbeta were shown to enter the complex at different ATP concentrations. Purified hMutLalpha showed an intrinsic affinity for DNA, with a preference for single-stranded over double-stranded DNA.
Plotz
Guido
G
Second Department of Medicine, Johann Wolfgang Goethe-University, Theodor Stern-Kai 7, D-60590 Frankfurt am Main, Germany.
Raedle
Jochen
J
Brieger
Angela
A
Trojan
Jörg
J
Zeuzem
Stefan
S
D048868:Adaptor Proteins, Signal Transducing; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D020137:Base Pair Mismatch; D015153:Blotting, Western; D002352:Carrier Proteins; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D006367:HeLa Cells; D006801:Humans; D046911:Macromolecular Substances; D008280:Magnetics; D008863:Microspheres; D000070976:Mismatch Repair Endonuclease PMS2; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D051718:MutS Homolog 2 Protein; D009363:Neoplasm Proteins / Q000378:metabolism*; D009687:Nuclear Proteins; D009994:Osmolar Concentration; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D013816:Thermodynamics
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C094443:G-T mismatch-binding protein; C501486:MLH1 protein, human; D046911:Macromolecular Substances; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; C088996:PMS1 protein, human; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; C094934:PMS2 protein, human; C497172:MSH2 protein, human; D000070976:Mismatch Repair Endonuclease PMS2; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D051718:MutS Homolog 2 Protein; D045643:DNA Repair Enzymes