title: Identification of determinants in the protein partners aCBF5 and aNOP10 necessary for the tRNA:Psi55-synthase and RNA-guided RNA:Psi-synthase activities.
abstract: Protein aNOP10 has an essential scaffolding function in H/ACA sRNPs and its interaction with the pseudouridine(Psi)-synthase aCBF5 is required for the RNA-guided RNA:Psi-synthase activity. Recently, aCBF5 was shown to catalyze the isomerization of U55 in tRNAs without the help of a guide sRNA. Here we show that the stable anchoring of aCBF5 to tRNAs relies on its PUA domain and the tRNA CCA sequence. Nonetheless, interaction of aNOP10 with aCBF5 can counterbalance the absence of the PUA domain or the CCA sequence and more generally helps the aCBF5 tRNA:Psi55-synthase activity. Whereas substitution of the aNOP10 residue Y14 by an alanine disturbs this activity, it only impairs mildly the RNA-guided activity. The opposite effect was observed for the aNOP10 variant H31A. Substitution K53A or R202A in aCBF5 impairs both the tRNA:Psi55-synthase and the RNA-guided RNA:Psi-synthase activities. Remarkably, the presence of aNOP10 compensates for the negative effect of these substitutions on the tRNA: Psi55-synthase activity. Substitution of the aCBF5 conserved residue H77 that is expected to extrude the targeted U residue in tRNA strongly affects the efficiency of U55 modification but has no major effect on the RNA-guided activity. This negative effect can also be compensated by the presence of aNOP10.
authors: Laboratoire de Maturation des ARN et Enzymologie Moléculaire, UMR 7567 CNRS-UHP, Nancy Université, Université Henri Poincaré-BP 239, 54506 Vandoeuvre-Lès-Nancy cedex, France.
authors: Sébastien
authors: S
authors: Muller
authors: Jean-Baptiste
authors: JB
authors: Fourmann
authors: Christine
authors: C
authors: Loegler
authors: Bruno
authors: B
authors: Charpentier
authors: Christiane
authors: C
authors: Branlant
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D019753:Intramolecular Lyases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019751:Intramolecular Transferases; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary; D011560:Pseudouridine / Q000378:metabolism; D043903:Pyrococcus abyssi / Q000235:genetics; D012343:RNA, Transfer / Q000737:chemistry* / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017384:Sequence Deletion; D014529:Uridine / Q000378:metabolism; D058727:RNA, Small Untranslated
chemical_list: D019843:Archaeal Proteins; D017411:Ribonucleoproteins, Small Nuclear; D011560:Pseudouridine; D012343:RNA, Transfer; D019751:Intramolecular Transferases; C096995:pseudouridine synthases; D019753:Intramolecular Lyases; D014529:Uridine; D058727:RNA, Small Untranslated
references: FEBS Lett. 1999 Nov 26;462(1-2):94-100
references: 10580099
references: RNA. 2004 Feb;10(2):231-9
references: 14730022
references: Biochemistry. 2000 Aug 8;39(31):9459-65
references: 10924141
references: Cell. 2001 Dec 28;107(7):929-39
references: 11779468
references: Cell. 2002 Apr 19;109(2):145-8
references: 12007400
references: Trends Biochem Sci. 2002 Jul;27(7):344-51
references: 12114023
references: Biochimie. 2002 Aug;84(8):775-90
references: 12457565
references: Nucleic Acids Res. 2003 Feb 1;31(3):869-77
references: 12560482
references: Curr Opin Struct Biol. 2003 Feb;13(1):49-55
references: 12581659
references: Proc Natl Acad Sci U S A. 2003 Oct 28;100(22):12648-53
references: 14566049
references: EMBO J. 2004 Apr 21;23(8):1857-67
references: 15044956
references: J Biol Chem. 2004 Apr 30;279(18):18107-10
references: 14999002
references: Structure. 2004 May;12(5):893-903
references: 15130481
references: RNA. 2004 Jul;10(7):1034-46
references: 15208440
references: J Biol Chem. 1988 Feb 15;263(5):2255-63
references: 3276686
references: Biochemistry. 1993 Sep 21;32(37):9754-62
references: 8373778
references: Biochemistry. 1995 Jul 11;34(27):8904-13
references: 7612632
references: RNA. 1995 Mar;1(1):102-12
references: 7489483
references: RNA. 1995 Jun;1(4):437-48
references: 7493321
references: Nucleic Acids Res. 1996 Jun 15;24(12):2411-5
references: 8710514
references: RNA. 1996 Nov;2(11):1079-93
references: 8903339
references: Biochemistry. 1998 Jan 6;37(1):339-43
references: 9425055
references: Biochemistry. 1998 Jan 6;37(1):344-51
references: 9425056
references: EMBO J. 1998 Oct 1;17(19):5783-95
references: 9755178
references: Nucleic Acids Res. 1999 Jan 1;27(1):196-7
references: 9847178
references: J Mol Evol. 1999 Mar;48(3):291-302
references: 10093218
references: RNA. 1999 Jun;5(6):751-63
references: 10376875
references: J Biol Chem. 1999 Jul 2;274(27):18880-6
references: 10383384
references: J Biol Chem. 1999 Aug 6;274(32):22225-30
references: 10428788
references: Mol Cell Biol. 1999 Nov;19(11):7461-72
references: 10523634
references: RNA. 2005 Feb;11(2):210-9
references: 15659360
references: J Biol Chem. 2005 Feb 25;280(8):6655-62
references: 15611063
references: J Mol Biol. 2005 May 6;348(3):563-73
references: 15826654
references: Genes Dev. 2005 May 15;19(10):1238-48
references: 15870259
references: Nucleic Acids Res. 2005;33(10):3133-44
references: 15933208
references: RNA. 2005 Sep;11(9):1420-9
references: 16120833
references: Biochemistry. 2005 Nov 29;44(47):15488-94
references: 16300397
references: Nat Struct Mol Biol. 2005 Dec;12(12):1101-7
references: 16286935
references: RNA. 2006 Jan;12(1):40-52
references: 16373493
references: Mol Cell. 2006 Jan 20;21(2):249-60
references: 16427014
references: Nucleic Acids Res. 2006;34(3):826-39
references: 16456033
references: Nature. 2006 Sep 21;443(7109):302-7
references: 16943774
references: Nucleic Acids Res. 2006;34(15):4293-301
references: 16920741
references: Nature. 2006 Oct 26;443(7114):956-60
references: 17051158
references: Chem Biol. 2006 Nov;13(11):1125-35
references: 17113994
references: Proc Natl Acad Sci U S A. 2007 Apr 17;104(16):6655-60
references: 17412831
references: Mol Cell. 2007 Apr 27;26(2):205-15
references: 17466623
references: Methods Enzymol. 2007;425:21-53
references: 17673078
references: Methods Enzymol. 2007;425:389-405
references: 17673092
references: Mol Cell. 2003 Feb;11(2):425-35
references: 12620230
references: Mol Microbiol. 2003 May;48(3):617-29
references: 12694609
references: RNA. 2003 Jun;9(6):711-21
references: 12756329
references: RNA. 2003 Jul;9(7):771-9
references: 12810910
references: RNA. 2003 Nov;9(11):1371-82
references: 14561887
references: Nucleic Acids Res. 2000 May 15;28(10):2075-83
references: 10773075