title: A functional interaction of SmpB with tmRNA for determination of the resuming point of trans-translation.
abstract: In trans-translation, transfer-messenger RNA (tmRNA), possessing a dual function as a tRNA and an mRNA, relieves a stalled translation on the ribosome with the help of SmpB. Here, we established an in vitro system using Escherichia coli translation and trans-translation factors to evaluate two steps of trans-translation, peptidyl transfer from peptidyl-tRNA to alanyl-tmRNA and translation of the resume codon on tmRNA. Using this system, the effects of several mutations upstream of the tag-encoding region on tmRNA were examined. These mutations affected translation of the resume codon rather than peptidyl transfer, and one of them, A84U/U85G, caused a shift of the resume codon by -1. We also found that U(85) is protected from chemical modification by SmpB. In the A84U/U85G mutant, the base of protection was shifted from 85 to 84. Another mutation, A86U, which caused a shift of the resume codon by +1, shifted the base of protection from 85 to 86. The protection at 85 was suppressed by a mutation in the tRNA-like domain critical to SmpB binding. These results suggest that SmpB serves to bridge two separate domains of tmRNA to determine the initial codon for tag-translation. A mutant SmpB with a truncation of the unstructured C-terminal tail failed to promote peptidyl transfer, although it still protected U(85) from chemical modification.
authors: Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University, Hirosaki 036-8561, Japan.
authors: Takayuki
authors: T
authors: Konno
authors: Daisuke
authors: D
authors: Kurita
authors: Kazuma
authors: K
authors: Takada
authors: Akira
authors: A
authors: Muto
authors: Hyouta
authors: H
authors: Himeno
mesh_terms: D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D012343:RNA, Transfer / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*
chemical_list: D012329:RNA, Bacterial; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C430328:small protein B; D012343:RNA, Transfer
references: EMBO J. 2000 Mar 1;19(5):1098-107
references: 10698950
references: EMBO J. 1999 Jul 1;18(13):3793-9
references: 10393194
references: FEBS Lett. 2000 Mar 31;470(3):345-9
references: 10745094
references: EMBO J. 2000 Jul 17;19(14):3762-9
references: 10899129
references: Genes Cells. 2000 Aug;5(8):627-35
references: 10947848
references: EMBO J. 2000 Dec 1;19(23):6612-21
references: 11101533
references: RNA. 2001 Jul;7(7):999-1012
references: 11453072
references: Nucleic Acids Res. 2001 Nov 15;29(22):4663-73
references: 11713316
references: J Mol Biol. 2001 Nov 16;314(1):9-21
references: 11724528
references: FEBS Lett. 2002 Mar 6;514(1):74-7
references: 11904185
references: Nucleic Acids Res. 2002 Apr 1;30(7):1620-9
references: 11917023
references: RNA. 1999 Aug;5(8):989-92
references: 10445873
references: Nucleic Acids Res. 1999 Sep 15;27(18):3667-75
references: 10471735
references: EMBO J. 1999 Oct 1;18(19):5423-33
references: 10508174
references: J Biol Chem. 2004 Dec 24;279(52):54202-9
references: 15494393
references: J Biol Chem. 2005 Feb 18;280(7):5503-9
references: 15596445
references: Proc Natl Acad Sci U S A. 2005 Feb 15;102(7):2316-21
references: 15699355
references: Nucleic Acids Res. 2005;33(8):2384-94
references: 15860775
references: Nucleic Acids Res. 2005;33(11):3529-39
references: 15972795
references: Mol Microbiol. 2005 Jul;57(2):565-75
references: 15978085
references: Nucleic Acids Res. 2005;33(17):5544-52
references: 16204455
references: Nucleic Acids Res. 2006 Jan 1;34(Database issue):D163-8
references: 16381838
references: J Biochem. 2005 Dec;138(6):729-39
references: 16428302
references: Nucleic Acids Res. 2006;34(6):1935-43
references: 16611927
references: Proc Natl Acad Sci U S A. 2006 Oct 31;103(44):16484-9
references: 17056712
references: J Biol Chem. 2007 Mar 2;282(9):6356-63
references: 17179154
references: RNA. 2007 Apr;13(4):503-10
references: 17299130
references: RNA. 2007 May;13(5):713-22
references: 17400816
references: Proc Natl Acad Sci U S A. 2007 May 15;104(20):8293-8
references: 17488812
references: Genes Cells. 2002 Mar;7(3):343-50
references: 11918677
references: EMBO J. 2002 Apr 2;21(7):1845-54
references: 11927568
references: Biochemistry. 2002 Jul 16;41(28):8826-36
references: 12102625
references: Eur J Biochem. 2003 Feb;270(3):463-75
references: 12542696
references: FEBS Lett. 2003 Jan 30;535(1-3):94-100
references: 12560085
references: J Bacteriol. 2003 Mar;185(6):1825-30
references: 12618446
references: J Biol Chem. 2003 Jul 25;278(30):27672-80
references: 12754221
references: Nature. 2003 Aug 7;424(6949):699-703
references: 12904796
references: Annu Rev Microbiol. 2003;57:101-23
references: 12730326
references: Nucleic Acids Res. 2004 Jan 1;32(Database issue):D104-8
references: 14681369
references: J Biol Chem. 2004 Jun 18;279(25):25978-85
references: 15069072
references: Nucleic Acids Res. 2004;32(14):4119-26
references: 15295039
references: J Mol Recognit. 1991 Jul-Dec;4(4):129-32
references: 1799462
references: Nucleic Acids Res. 1994 Aug 25;22(16):3392-6
references: 7521527
references: Proc Natl Acad Sci U S A. 1994 Sep 27;91(20):9223-7
references: 7524073
references: Science. 1996 Feb 16;271(5251):990-3
references: 8584937
references: RNA. 1996 Dec;2(12):1306-10
references: 8972778
references: RNA. 1997 Jan;3(1):89-103
references: 8990402
references: J Mol Biol. 1997 May 23;268(5):803-8
references: 9180372
references: Trends Biochem Sci. 1998 Jan;23(1):25-9
references: 9478132
references: J Mol Biol. 1999 Feb 26;286(3):733-44
references: 10024447
references: Biochemistry. 2000 Mar 14;39(10):2652-8
references: 10704215