Isolation and characterization of a dominant negative mutant of Bacillus subtilis GTP-binding protein, YlqF, essential for biogenesis and maintenance of the 50 S ribosomal subunit.
The circularly permuted GTPase YlqF is essential for cell viability and is broadly conserved from Gram-positive bacteria to eukaryotes. We previously reported that YlqF participates in the late step of 50 S ribosomal subunit assembly in Bacillus subtilis. Here, we demonstrate that an N-terminal deletion mutant of YlqF (YlqFDeltaN10) inhibits cell growth even in the presence of wild-type YlqF. In contrast to the wild-type protein, the GTPase activity of this mutant was not stimulated by the 50 S subunit and did not dissociate from the premature 50 S subunit. Thus, YlqFDeltaN10 acts as a competitive inhibitor of wild-type YlqF. Premature 50 S subunit lacking ribosomal protein L27 and with a reduced amount of L16 accumulated in YlqFDeltaN10-overexpressing cells and in YlqF-depleted cells, suggesting that YlqFDeltaN10 binds to the premature 50 S subunit. Moreover, premature 50 S subunit from both YlqFDeltaN10-overexpressing and YlqF-depleted cells more strongly enhanced the GTPase activity of YlqF than the mature 50 S subunit of the 70 S ribosome. Collectively, our results indicate that YlqF is targeted to the premature 50 S subunit lacking ribosomal proteins L16 and L27 to assemble functional 50 S subunit through a GTPase activity-dependent conformational change of 23 S rRNA.
Matsuo
Yoshitaka
Y
Department of Bioinformatics and Genomics, Graduate School of Information Science, Nara Institute of Science and Technology, 8916-5, Takayama, Ikoma, Nara 630-0101, Japan.
Oshima
Taku
T
Loh
Pek Chin
PC
Morimoto
Takuya
T
Ogasawara
Naotake
N
D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005798:Genes, Bacterial; D005799:Genes, Dominant; D009154:Mutation; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D012338:RNA, Ribosomal, 23S / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D012329:RNA, Bacterial; D012338:RNA, Ribosomal, 23S; D012269:Ribosomal Proteins; C027937:ribosomal protein L16; C034661:ribosomal proteins L27; D019204:GTP-Binding Proteins