Use of selectively trypsinized nucleosome core particles to analyze the role of the histone "tails" in the stabilization of the nucleosome.
Using immobilized trypsin and an appropriate fractionation procedure, we have been able to prepare, for the first time, nucleosome core particles containing selectively trypsinized histone domains. The particles thus obtained: [(H3T-H4T)2-2(H2AT-H2BT)].DNA; [(H3-H4)2-2(H2AT-H2BT)].DNA; [H3T-H4T)2-2(H2A-H2B)].DNA (where T means trypsinized), together with the non-trypsinized controls have been characterized using the following techniques: analytical ultracentrifugation, circular dichroism, thermal denaturation and DNAse I digestion. The major aim of this study was to analyze the role of the amino-terminal regions (the histone "tails") on the stability of the nucleosome in solution. The data obtained from this analysis clearly show that stability of the nucleosome core particle to dissociation (below a salt concentration of 0.7 M-NaCl) is not affected by the presence or the absence of any of the N-terminal regions of the histones. Furthermore, these histone regions make very little contribution, if any, to the conformational transition that nucleosomes undergo in this range of salt concentrations. They play, however, a very important role in determining the thermal stability of the particle, as reflected in the dramatic alterations exhibited by the melting profiles upon selective removal of these tails by trypsinization. The melting data can be explained by a simple hypothesis that ascribes interaction of H2A/H2B and H3/H4 tails to particular regions of the nucleosomal DNA.
Ausio
J
J
Department of Biochemistry and Biophysics, Oregon State University, Corvallis 97331-6503.
Dong
F
F
van Holde
K E
KE
D000818:Animals; D002645:Chickens; D002942:Circular Dichroism; D003850:Deoxyribonuclease I / Q000378:metabolism; D004912:Erythrocytes; D006657:Histones / Q000502:physiology*; D006358:Hot Temperature; D009707:Nucleosomes / Q000378:metabolism*; D011489:Protein Denaturation; D013816:Thermodynamics; D014357:Trypsin / Q000378:metabolism*
D006657:Histones; D009707:Nucleosomes; D003850:Deoxyribonuclease I; D014357:Trypsin