Binding of 16S rRNA to chloroplast 30S ribosomal proteins blotted on nitrocellulose

Abstract

Protein-RNA associations were studied by a method using proteins blotted on a nitrocellulose sheet. This method was assayed with Escherichia Coli 30S ribosomal components. In stringent conditions (300 mM NaCl or 20 degrees C) only 9 E. coli ribosomal proteins strongly bound to the 16S rRNA: S4, S5, S7, S9, S12, S13, S14, S19, S20. 8 of these proteins have been previously found to bind independently to the 16S rRNA. The same method was applied to determine protein-RNA interactions in spinach chloroplast 30S ribosomal subunits. A set of only 7 proteins was bound to chloroplast rRNA in stringent conditions: chloroplast S6, S10, S11, S14, S15, S17 and S22. They also bound to E. coli 16S rRNA. This set includes 4 chloroplast-synthesized proteins: S6, S11, S15 and S22. The core particles obtained after treatment by LiCl of chloroplast 30S ribosomal subunit contained 3 proteins (S6, S10 and S14) which are included in the set of 7 binding proteins. This set of proteins probably play a part in the early steps of the assembly of the chloroplast 30S ribosomal subunit.

Chloroplast ribosomes and protein synthesis.

“…The main observations on land plants and C. reinhardtii are as follows. (i) Seven chloroplast ribosomal proteins, four of which are made in the chloroplast, bind to chloroplast or E. coli 16S rRNA, in agreement with the seven E. coli ribosomal proteins known to bind to 16S rRNA (531). (ii) Two 5S rRNA-binding proteins have been detected in spinach (L22 and "CS-12" [651]), in contrast to three in E. coli (L5, L18, L25).…”

The components of the plastid ribosome are not accumulated synchronously during the early development of spinach plants

“…4B) could also be primary RNAbinding proteins which are important for the early assembly of the ribosomal particles [40]. This may be the case at least for r-proteins CS-S10 and CS-SI4 which have been identified as being primary 16S rRNA binding proteins in spinach [25].…”

Immunoblotting and dot blotting

“…Proteins that bind to defined sequences of DNA or RNA can also be studied by protein blotting, which provides a promising way of identifying putative gene regulatory proteins. Nucleic acid binding proteins from HeLa ceils, viruses, oocytes, Drosophila and heat shock proteins have been analysed by blotting and overlay with labelled DNA, RNA or histones (Bowen et al, 1980;Baumann and Hand, 1982;Aubertin et al, 1983;Richter and Smith, 1983;Gabor and Bennett, 1984;Rozier and Mache, 1984;Wolff et al, 1985). Miskimins et al (1985) used a DNA probe to identify nuclear proteins that bind to the promoter region of the transferrin receptor gene.…”