Isolation, crystallization, and investigation of ribosomal protein S8 complexed with specific fragments of rRNA of bacterial or archaeal origin.
The core ribosomal protein S8 binds to the central domain of 16S rRNA independently of other ribosomal proteins and is required for assembling the 30S subunit. It has been shown with E. coli ribosomes that a short rRNA fragment restricted by nucleotides 588-602 and 636-651 is sufficient for strong and specific protein S8 binding. In this work, we studied the complexes formed by ribosomal protein S8 from Thermus thermophilus and Methanococcus jannaschii with short rRNA fragments isolated from the same organisms. The dissociation constants of the complexes of protein S8 with rRNA fragments were determined. Based on the results of binding experiments, rRNA fragments of different length were designed and synthesized in preparative amounts in vitro using T7 RNA-polymerase. Stable S8-RNA complexes were crystallized. Crystals were obtained both for homologous bacterial and archaeal complexes and for hybrid complexes of archaeal protein with bacterial rRNA. Crystals of the complex of protein S8 from M. jannaschii with the 37-nucleotide rRNA fragment from the same organism suitable for X-ray analysis were obtained.
Tishchenko
S V
SV
Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region, 142290 Russia. sveta@vega.protres.ru
Vassilieva
J M
JM
Platonova
O B
OB
Serganov
A A
AA
Fomenkova
N P
NP
Mudrik
E S
ES
Piendl
W
W
Ehresmann
C
C
Ehresmann
B
B
Garber
M B
MB
D001665:Binding Sites; D003460:Crystallization; D008274:Magnesium / Q000737:chemistry / Q000378:metabolism; D017017:Methanococcus / Q000737:chemistry / Q000235:genetics; D009690:Nucleic Acid Conformation; D019642:RNA, Archaeal / Q000737:chemistry / Q000378:metabolism; D012329:RNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D012335:RNA, Ribosomal / Q000737:chemistry* / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D016963:Thermus thermophilus / Q000737:chemistry / Q000235:genetics
D019642:RNA, Archaeal; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012269:Ribosomal Proteins; C030254:ribosomal protein S8; D008274:Magnesium