Date of Award
Doctor of Philosophy
Biochemistry, Biophysics and Molecular Biology
For an understanding of the molecular events that occur on the ribosome during protein biosynthesis, the interaction of proteins and RNA within the ribosome must be understood. In this study, the interaction of ribosomal RNAs with ribosomal proteins was studied using an affinity chromatography procedure. Immobilized RNA columns were prepared by first oxidizing the 3' terminus of the RNA and then covalently coupling this to dihydrazide-Sepharose. In some experiments, columns were prepared in which RNA was attached to dihydrazide-Sepharose through its 5' terminus. Initial experiments investigated the interaction of 3'-bound E. coli 5S RNA with E.coli 50S ribosomal proteins. Proteins were bound to the immobilized RNA at low salt concentration and then eluted by high salt. Using buffer containing 0.3M KCl, proteins L5, L18 and L25 were bound to E. coli 5S RNA. At 0.2M KCl, proteins L2 and L17 were also bound but with less affinity than proteins L5, L18 and L25. The interaction of E. coli 50S ribosomal proteins with yeast 5S RNA and yeast 5.8S RNA was also studied, in an attempt to identify the eukaryotic RNA which might be the functional analog of E. coli 5S RNA. However, no proteins were bound to either yeast RNA at 0.3M KCl. At 0.2M KCl proteins L2 and L17 were bound to both yeast RNAs. Proteins L2 and L17 were also bound to columns of immobilized poly (U), but failed to bind to columns containing oligo (U)(,6) or Sepharose which lacked RNA. Using columns containing 5'-immobilized RNA, proteins L2 and L17 were bound to E. coli 5S RNA, yeast 5S RNA and 5.8S RNA, but proteins L5, L18 and L25 were bound only to E. coli 5S RNA;The interaction of yeast 60S ribosomal proteins and rat liver of 60S ribosomal proteins with E. coli 5S RNA, yeast 5S RNA and yeast 5.8S RNA was studied at several salt concentrations. At 0.2 M KCl, yeast 60S proteins L2/3, L10, L21 and L23 bound to all 3 RNAs, while protein L30 bound to yeast 5S RNA and L12 bound to yeast 5.8S RNA. Yeast RNAs were also able to bind significant amounts of rat liver 60S proteins. At 0.3M KCl, about 1/3 of the 44 rat liver 60S proteins were bound. Only 4 major proteins, L3, L4, L6 and L35 were bound to yeast 5.8S RNA at 0.45M KCl;The binding of small subunit ribosomal proteins from E. coli and yeast to yeast 5S RNA and yeast 5.8S RNA was studied and a small number of proteins were bound to both RNAs. E. coli 30S proteins bound were identified as S4, S9 and S28; with S17 the major yeast 40S protein bound.
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/
Janice Ann Kolberg
Kolberg, Janice Ann, "Studies on the interactions of low molecular weight ribosomal RNAs and ribosomal proteins " (1984). Retrospective Theses and Dissertations. 7774.