Site-specific proteolytic cleavage of Ku protein bound to DNA.
Ku protein, a relatively abundant nuclear protein associated with DNA of mammalian cells, is known to be a heterodimer with subunits of 85 and 72 kDa which binds in vitro to DNA ends and subsequently translocates along the molecule. The functional role played by this protein in the cell, however, remains to be elucidated. We have observed here that Ku protein, purified from cultured monkey cells, is the target of specific endoproteolysis in vitro, by which the 85 kDa subunit is cleaved at a precise site while the 72 kDa subunit remains intact. This cleavage releases an 18 kDa polypeptide and converts Ku protein into a heterodimer composed of the 72 kDa subunit associated with a 69 kDa fragment from the 85 kDa subunit. The proteolyzed form of Ku protein, denoted Ku', has DNA binding properties similar to those of Ku protein. The proteolytic mechanism, which is inhibited by leupeptin and chymostatin, is extremely sensitive to ionic conditions, in particular to pH, being very active at pH 7.0 and completely inhibited at pH 8.0. In addition, cleavage occurs only when Ku protein is bound to DNA, not free in solution. We suggest that in vivo, such proteolysis might be necessary for Ku protein function at some stage of the cell cycle.
Paillard
S
S
Institut Jacques Monod, Paris, France.
Strauss
F
F
D000818:Animals; D034961:Antigens, Nuclear; D001665:Binding Sites; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D004265:DNA Helicases; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D000072200:Ku Autoantigen; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism; D011487:Protein Conformation
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D010447:Peptide Hydrolases; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen