Fractionation of constituents of ribonucleoproteins containing heterogeneous nuclear ribonucleic acid

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Abstract
A method of fractionation of hnRNP [heterogeneous nuclear ribonucleoprotein] constituents adaptable to large-scale preparation from rat brains is presented. It is based on differential resistance to salt dissociation of the 2 classes of units of hnRNP, the 30-50S monoparticles and the heterogeneous complexes. The monoparticle proteins were released from hnRNP by 0.4 M NaCl. They were separated from the salt-resistant RNP corresponding to the heterogeneous complexes in 3 steps: chromatography on DEAE-cellulose, high-speed centrifugation and Bio-Gel chromatography. The latter chromatography permitted a 1st fractionation of monoparticle proteins according to molecular weight. Such fractions may serve for purification of individual proteins of MW below 80,000. After the 2 first steps, 2 fractions of salt-resistant RNP were obtained. In addition to heterogeneous RNA up to 30S, small nuclear RNA were detected which represented 6% of total RNA. The protein pattern was complex, and no clear-cut segregation of groups of proteins could be observed between the 2 fractions. They were both highly enriched in phosphoproteins as compared to monoparticle proteins. In another fraction corresponding to the void volume of Bio-Gel chromatography, 1/3 of the RNA was small nuclear RNA. This fraction contains snRNP [small nuclear ribonucleoprotein] in addition to free proteins of MW > 80,000 and to salt-resistant RNP similar to those described above but of small size.