Analysis of the binding of high mobility group protein 17 to the nucleosome core particle by 1H NMR spectroscopy.
The binding of high mobility group (HMG) protein 17 to the nucleosome core particle has been studied in D2O solution using 1H NMR at 500 MHz. Spectra were obtained for purified HMG 17, purified nucleosome core particles, and the reconstituted HMG 17-nucleosome core particle complex at 0.1, 0.2, 0.3, and 0.4 M NaCl. Subtraction of the core particle spectra from spectra of the core particle reconstituted with HMG 17 demonstrated those regions of HMG 17 which interact with the nucleosome at different ionic strengths; the resonance peaks of interacting groups are broadened due to their restricted mobility. At 0.1 M NaCl, the mobility of all the amino acid side chains of HMG 17 was restricted, indicating complete binding of HMG 17 to the much larger nucleosome core particle. At 0.2 M NaCl most of the amino acids were free with the exception of arginine and proline which are confined to or predominant in the basic central region of HMG 17. These amino acids were completely free only at 0.4 M NaCl. We conclude that the entire HMG 17 molecule interacts with the nucleosome core particle at physiological ionic strength. The acidic COOH-terminal region of HMG 17 is released from interaction with the core histones at an NaCl concentration between 0.1 and 0.2 M and so binds weakly at physiological ionic strength. The basic central region binds more strongly to the core particle DNA, being completely released only at much higher ionic strength, between 0.3 and 0.4 M NaCl.
Cook
G R
GR
Department of Biological Chemistry, School of Medicine, University of California, Davis 95616.
Minch
M
M
Schroth
G P
GP
Bradbury
E M
EM
D000595:Amino Acid Sequence; D000818:Animals; D006609:High Mobility Group Proteins / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009707:Nucleosomes / Q000378:metabolism*; D009994:Osmolar Concentration; D012756:Sheep
D006609:High Mobility Group Proteins; D009707:Nucleosomes