A complete set of RNA-binding proteins was isolated from the ribosome-free extract of rabbit reticulocytes using the method of affinity chromatography on RNA covalently coupled with Sepharose. The purity of the isolated proteins was no less than 90%. These proteins comprised about 1 % of the total protein of the extract and included the main polypeptide chains of three sizes, with molecular weights of about 95000, 49000 and 36000, as well as numerous minor components. An analogous set of proteins was observed as a result of chromatography of the extract on the column with poly(U) covalently coupled with Sepharose. The protein with the molecular weight of 49000 had the highest affinity to RNA.A special type of ribonucleoprotein particles denoted as informosomes were revealed in cytoplasmic extracts of eukaryotic cells (fish embryos) in 1964 [I] (see also review [2]). These particles contained messenger RNA and differed from other cellular ribonucleoproteins by their comparably low buoyant density in CsCl (about 1.4 g/cm3). Similar particles were later found in nuclear extracts [3] and within polyribosomes [4-61. A special class of proteins capable of interacting with high-molecular-weight RNA and forming ribonucleoprotein particles similar to informosomes were also found in cytoplasmic [7,8] and nuclear [9] extracts of animal cells; it was speculated that these informosome-forming (RNA-binding) proteins represent excess informosomal proteins in a free state [lo].Numerous attempts have been made recently to elucidate the functions of informosomal proteins. The first step in solving this problem is the isolation of pure informosomes and RNA-binding proteins. To isolate the latter, affinity chromatography on RNA or synthetic polyribonucleotides covalently bound with a solid matrix was used [ l l -141.However, in the majority of previous reports on affinity isolation of RNA-binding proteins neither the completeness of the isolated sets of these proteins nor their purity was controlled in a sufficient degree. Contradictory results were obtained concerning the dependence of the isolated protein sets on the type of matrix-bound polyribonucleotide used.

No abstract

No abstract

The complete set of rabbit reticulocyte PNA-binding proteins was compared with the proteins of rabbit reticulocyte polyribosomal messenger ribonucleoproteins as well as with the proteins of rat liver nuclear 30-S ribonucleoprotein particles, using electrophoresis in polyacrylamide gel in the presence of sodium dodecylsulfate. The purity of the two ribonucleoprotein preparations was 90 % and 95 %, respectively. Rabbit reticulocyte RNA-binding proteins included main polypeptide chains of three sizes with molecular weights of about 95000, 49000 and 36000. Rabbit reticulocyte polyribosomal messenger ribonucleoproteins with a sedimentation coefficient of about 13 S and a buoyant density of 1.48 g/cm3 in CsCl were shown to contain polypeptide chains of four sizes with molecular weights of about 78000,49000,46000 and 33000. Rat liver nuclear 30-S ribonucleoprotein particles with a buoyant density of 1.41 -1.43 g/cm3 in CsCl contained polypeptide chains of two sizes with molecular weights of about 40000 and 37000. A parallel electrophoresis of the RNAbinding proteins and proteins of the ribonucleoprotein particles as well as the electrophoresis of their mixtures showed that one of the main RNA-binding proteins with a molecular weight of 49000 is indistinguishable from the corresponding protein of polyribosomal messenger ribonucleoproteins. The two other RNA-binding proteins could not be detected in the polyribosomal messenger ribonucleoproteins. The polypeptides contained in nuclear 30-S ribonucleoprotein particles were found neither in polyribosomal messenger ribonucleoproteins nor among RNA-binding proteins.In eukaryotic cells all messenger RNA is complexed with protein and thus present in the form of ribonucleoprotein particles characterized by a comparatively low buoyant density in CsCl (about 1.4 g/cm3). The term informosomes was suggested to denote these particles [l, 21. Three classes of such particles are distinguished: nuclear ribonucleoproteins [3,4], free cytoplasmic informosomes [l, 2,5] and polyribosomal messenger ribonucleoproteins 16 -81. Together with informosomes a special fraction of RNA-binding proteins is also present in the cytoplasm [9] and nucleus [lo]; these proteins are capable of interacting with RNA and forming artificial ribonucleoprotein particles similar to informosomes in their sedimentation coefficients and buoyant density values.Many studies in the last few years have been performed to determine protein compositions of the particles of the different classes and to compare them with each other. However, most of them can be regarded as only preliminary due to the absence of appropriate guarantees of purity of the preparations analyzed from protein contamination. Here we propose a method of evaluating protein contamination in ribonucleoprotein particle preparations and compare the complete set of cytoplasmic RNA-binding proteins of rabbit reticulocytes with the proteins of polyribosoinal messenger ribonucleoproteins (90% protein purity) of the same cells, on the one hand, and rat liver nuc...