Fluctuations of transfer RNAs between classical and hybrid states.
Adjacent transfer RNAs (tRNAs) in the A- and P-sites of the ribosome are in dynamic equilibrium between two different conformations called classical and hybrid states before translocation. Here, we have used single-molecule fluorescence resonance energy transfer to study the effect of Mg(2+) on tRNA dynamics with and without an acetyl group on the A-site tRNA. When the A-site tRNA is not acetylated, tRNA dynamics do not depend on [Mg(2+)], indicating that the relative positions of the substrates for peptide-bond formation are not affected by Mg(2+). In sharp contrast, when the A-site tRNA is acetylated, Mg(2+) lengthens the lifetime of the classical state but does not change the lifetime of the hybrid state. Based on these findings, the classical state resembles a state with direct stabilization of tertiary structure by Mg(2+) ions whereas the hybrid state resembles a state with little Mg(2+)-assisted stabilization. The antibiotic viomycin, a translocation inhibitor, suppresses tRNA dynamics, suggesting that the enhanced fluctuations of tRNAs after peptide-bond formation drive spontaneous attempts at translocation by the ribosome.
Kim
Harold D
HD
Department of Physics, Stanford University, Stanford, California 94305, USA.
Puglisi
Joseph D
JD
Chu
Steven
S
D001703:Biophysics / Q000379:methods; D031541:Fluorescence Resonance Energy Transfer; D007477:Ions; D008274:Magnesium / Q000737:chemistry*; D008956:Models, Chemical; D009690:Nucleic Acid Conformation; D010441:Peptide Chain Elongation, Translational; D010455:Peptides / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D021381:Protein Transport; D012343:RNA, Transfer / Q000737:chemistry*; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry*; D012270:Ribosomes / Q000737:chemistry*; D013997:Time Factors; D014756:Viomycin / Q000737:chemistry / Q000494:pharmacology*
D007477:Ions; D010455:Peptides; D012346:RNA, Transfer, Amino Acyl; D011092:Polyethylene Glycols; D012343:RNA, Transfer; D008274:Magnesium; D014756:Viomycin
Nucleic Acids Res. 2005;33(5):1618-27
15767286
J Am Chem Soc. 2005 Mar 23;127(11):3801-6
15771514
Phys Rev Lett. 2005 Mar 18;94(10):108101
15783528
Cell. 2006 Apr 7;125(1):71-84
16615891
Nature. 2007 Mar 22;446(7134):454-7
17377584
Nature. 2000 Jul 20;406(6793):318-22
10917535
J Cell Biol. 2000 Aug 7;150(3):447-60
10931859
Nat Struct Biol. 2000 Nov;7(11):1027-31
11062557
J Mol Biol. 2000 Dec 15;304(5):707-13
11124020
Science. 2001 May 4;292(5518):883-96
11283358
Nat Struct Biol. 2002 Mar;9(3):225-30
11828326
Proc Natl Acad Sci U S A. 2002 Apr 2;99(7):4284-9
11929999
Mol Cell. 2002 May;9(5):1125-31
12049747
Proc Natl Acad Sci U S A. 2002 Sep 3;99(18):11670-5
12185246
J Mol Biol. 2003 Mar 21;327(2):369-81
12628244
Science. 2003 May 16;300(5622):1159-62
12750524
Cell. 2003 Jul 11;114(1):123-34
12859903
Proc Natl Acad Sci U S A. 2003 Aug 5;100(16):9302-7
12869691
Proc Natl Acad Sci U S A. 2003 Aug 5;100(16):9319-23
12878726
Proc Natl Acad Sci U S A. 2004 May 25;101(21):7897-901
15141076
Biochemistry. 2004 Jun 15;43(23):7575-83
15182199
Proc Natl Acad Sci U S A. 2004 Aug 31;101(35):12893-8
15317937
J Struct Biol. 2004 Sep;147(3):302-14
15450299
J Mol Biol. 2004 Nov 5;343(5):1183-94
15491605
Eur J Biochem. 1977 Dec;81(3):491-7
202460
Mol Gen Genet. 1978 May 31;161(3):261-5
209306
Eur J Biochem. 1986 Dec 15;161(3):715-21
3024981
Biochimie. 1987 Aug;69(8):879-84
3122849
Cell. 1989 May 19;57(4):585-97
2470511
Nature. 1989 Nov 9;342(6246):142-8
2682263
Nat Struct Mol Biol. 2004 Nov;11(11):1101-6
15475967