Independent in vitro assembly of all three major morphological parts of the 30S ribosomal subunit of Thermus thermophilus.
Fragments of the 16S rRNA of Thermus thermophilus representing the 3' domain (nucleotides 890-1515) and the 5' domain (nucleotides 1-539) have been prepared by transcription in vitro. Incubation of these fragments with total 30S ribosomal proteins of T. thermophilus resulted in formation of specific RNPs. The particle assembled on the 3' RNA domain contained seven proteins corresponding to Escherichia coli ribosomal proteins S3, S7, S9, S10, S13, S14, and S19. All of them have previously been shown to interact with the 3' domain of the 16S RNA and to be localized in the head of the 30S ribosomal subunit. The particle formed on the 5' RNA domain contained five ribosomal proteins corresponding to E. coli proteins S4, S12, S17, S16, and S20. These proteins are known to be localized in the main part of the body of the 30S subunit. Both types of particle were compact and had sedimentation coefficients of 15.5 S and 13 S, respectively. Together with our recent demonstration of the reconstitution of the RNA particle representing the platform of the T. thermophilus 30S ribosomal subunit [Agalarov, S.C., Zheleznyakova, E.N., Selivanova, O.M., Zheleznaya, L.A., Matvienko, N.I., Vasiliev, V.D. & Spirin, A.S. (1998) Proc. Natl Acad. Sci. USA 95, 999-1003], these experiments establish that all three main structural lobes of the small ribosomal subunit can be reconstituted independently of each other and prepared in the individual state.
Agalarov
S C
SC
Laboratory of Mechanisms of Protein Biosynthesis, Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region, Russia.
Selivanova
O M
OM
Zheleznyakova
E N
EN
Zheleznaya
L A
LA
Matvienko
N I
NI
Spirin
A S
AS
D002499:Centrifugation, Density Gradient; D015180:Electrophoresis, Gel, Two-Dimensional; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D012269:Ribosomal Proteins / Q000737:chemistry*; D013395:Sucrose / Q000494:pharmacology; D016963:Thermus thermophilus / Q000737:chemistry* / Q000378:metabolism; D014158:Transcription, Genetic
D012269:Ribosomal Proteins; C040469:ribosomal protein S30; D013395:Sucrose; D012313:RNA