Fractionation of the nuclear matrix. I. Partial separation into matrix protein fibrils and a residual ribonucleoprotein fraction.
Isolated rat liver nuclear matrices have been partially separated by means of mild sonication into a matrix protein (matricin) fraction and a residual ribonucleoprotein (RNP) fraction. The initial matricin fraction is composed largely of protein (91.1%) but also contains significant amounts of DNA (8.4%). Reconstruction experiments indicate that this DNA is not the result of the artifactual binding of DNA to the matrix during the extraction procedures. Subsequent treatment with DNase I results in purified matricin composed of greater than 99.5% protein. SDS acrylamide gel electrophoresis of the matrix protein fibrils reveals only three bands: the primary matrix polypeptides of 62,000, 66,000, and 70,000 daltons. Electron microscopy demonstrates a diffuse reticulum with fibrils as thin as 30--50 A and the presence of 80--100-A globular structures. The residual RNP fraction is composed largely of protein (80.1%) and RNA (19.5%), with only traces of DNA (1.1%). Over 98% of the total matrix-associated RNA is recovered in this fraction. SDS acrylamide gel electrophoresis indicates an enrichment in both low and high molecular weight secondary matrix polypeptides, although the 60,000--70,000-dalton polypeptides are present in significant amounts as well. Ultrastructural analysis of the residual RNP fraction reveals distinct electron-dense-staining matrix particles (150--350 A) attached to a fibrous matricin network.
D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002467:Cell Nucleus / Q000648:ultrastructure*; D002868:Chromosomal Proteins, Non-Histone / Q000302:isolation & purification*; D003852:Deoxyribonucleoproteins / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000648:ultrastructure; D008970:Molecular Weight; D009698:Nucleoproteins / Q000302:isolation & purification*; D051381:Rats; D012261:Ribonucleoproteins / Q000302:isolation & purification*
D002868:Chromosomal Proteins, Non-Histone; D003852:Deoxyribonucleoproteins; D009698:Nucleoproteins; D012261:Ribonucleoproteins