Functional investigation of residue G791 of Escherichia coli 16S rRNA: implication of initiation factor 1 in the restoration of P-site function.
Using a specialized ribosome system, previous studies have identified G791 in Escherichia coli 16S rRNA as an invariant and essential residue for ribosome function. To investigate the functional role of G791, we searched for multicopy suppressors that partially restored the protein synthesis ability of mutant ribosomes bearing a G to U substitution at position 791 (U791 ribosomes). Analyses of isolated multicopy suppressors showed that overexpression of initiation factor 1 (IF1) enhanced the protein synthesis ability of U791 ribosomes. In contrast, overexpression of initiation factor 2 (IF2) or IF3 did not enhance the protein synthesis ability of wild-type or U791 ribosomes, and overexpression of IF1 did not affect the function of wild-type or mutant ribosomes bearing nucleotide substitutions in other regions of 16S rRNA. Analyses of sucrose gradient profiles of ribosomes showed that overexpression of IF1 marginally enhanced the subunit association of U791 ribosomes and indicated lower binding affinity of U791 ribosomes to IF1. Our findings suggest the involvement of IF1 in the restoration of the P-site function that was impaired by a nucleotide substitution at residue G791.
Department of Life Sciences (BK21 program), Chung-Ang University, Seoul, Korea.
Woo-Seok
WS
Song
Sang-Mi
SM
Ryou
Hong-Man
HM
Kim
Che Ok
CO
Jeon
Jong-Myung
JM
Kim
Seung Hyun
SH
Han
Si Wouk
SW
Kim
Jin P
JP
Szatkiewicz
Philip R
PR
Cunningham
Kangseok
K
Lee
D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D039861:Prokaryotic Initiation Factor-1 / Q000235:genetics / Q000378:metabolism*; D039881:Prokaryotic Initiation Factor-2 / Q000235:genetics; D039665:Prokaryotic Initiation Factor-3 / Q000235:genetics; D014176:Protein Biosynthesis; D012336:RNA, Ribosomal, 16S / Q000235:genetics / Q000378:metabolism*; D054657:Ribosome Subunits / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism*; D013489:Suppression, Genetic
D039861:Prokaryotic Initiation Factor-1; D039881:Prokaryotic Initiation Factor-2; D039665:Prokaryotic Initiation Factor-3; D012336:RNA, Ribosomal, 16S