Mg2+-dependent unwinding of DNA by nonhistone chromosomal protein HMG(1 + 2) from pig thymus as determined by DNA melting temperature analysis.
In the previous studies with endonucleases specific for single-stranded DNA, we have indicated that the nonhistone chromosomal protein HMG(1 + 2) prepared from pig thymus has an activity to unwind DNA partially at low protein-to-DNA weight ratios (Yoshida, M. & Shimura, K. (1984) J. Biochem. 95, 117-124). In the present work, we have pursued the unwinding reaction by HMG(1 + 2) by thermal melting temperature analysis of DNA, and by investigating the effect of Mg2+ on the reaction. The melting temperature of DNA in the presence of HMG(1 + 2) at low protein weight ratios decreased in 2 mM Tris-HCl, pH 7.8, whereas it increased at higher ratios. The depressions of melting temperature by HMG(1 + 2) at low ratios were not observed either in the system of 2 mM Tris-HCl, pH 7.8, containing EDTA or in the system containing samples treated in advance with EDTA. An addition of Mg2+ to the system reproduced the depression of melting temperature at low protein-to-DNA ratios as well as the increase at higher ratios. Analysis by Mg2+-equilibrated gel filtration revealed that HMG(1 + 2) is a Mg2+-binding protein. However, the depression of melting temperature at low protein-to-DNA ratios was not due to removal of Mg2+ from DNA by HMG(1 + 2). From these results, it is concluded that HMG(1 + 2) causes a partial DNA unwinding detectable by thermal melting temperature analysis of DNA, and that Mg2+ is necessary for the unwinding reaction.
D000818:Animals; D002850:Chromatography, Gel; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D002875:Chromosomes / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D006609:High Mobility Group Proteins; D008274:Magnesium / Q000378:metabolism / Q000502:physiology*; D009691:Nucleic Acid Denaturation; D013552:Swine; D013816:Thermodynamics; D013950:Thymus Gland
D002868:Chromosomal Proteins, Non-Histone; D006609:High Mobility Group Proteins; D004247:DNA; D008274:Magnesium