ライフサイエンスコーパス: Met-tRNA

1 bound to the 40S preinitiation complex (40S.Met-tRNA(i).eIF2.GTP), promoted by eIF5, must occur only

2 nhibits the eIF3-dependent conversion of 40S/Met-tRNA(i)(Met)/mRNA to translationally competent 80S/M

3 )(Met)/mRNA to translationally competent 80S/Met-tRNA(i)(Met)/mRNA initiation complexes to repress co

4 r complex containing eIFs 1, 2, 3, and 5 and Met-tRNA(i)(Met), whose formation is required for an opt

5 tifactor complex (MFC) with eIFs 1, 2, 5 and Met-tRNA(i)(Met).

6 ation with eIF3 promotes binding of eIF2 and Met-tRNA(i)(Met) to 40S ribosomes.

7 ernary complex (TC) comprising eIF2, GTP and Met-tRNA(i).

8 at facilitates ribosomal subunit joining and Met-tRNA(i) binding to ribosomes in all three kingdoms o

9 of eIF2gamma that bind the alpha-subunit and Met-tRNA(i)(Met), respectively.

10 Gly-tRNA, Ala-tRNA, Ser-tRNA, Pro-tRNA, and Met-tRNA.

11 n of acceptor proteins using Leu-, Phe-, and Met-tRNAs as amino acid donors.

12 g in vitro binding of initiator Met-tRNA (as Met-tRNA(i).eIF2.GTP ternary complex) to 40 S ribosomal

13 ith the 40S initiation complex (40S*eIF3*AUG*Met-tRNA(f)*eIF2*GTP) and, acting as a GTPase activating

14 ith the 40S initiation complex (40S-eIF3-AUG-Met-tRNA(f)-eIF2-GTP) to promote the hydrolysis of ribos

15 S ribosomal initiation complex (40S.eIF3.AUG.Met-tRNA(f).eIF2.GTP) to promote the hydrolysis of bound

16 pendent of the canonical initiator tRNA (AUG/Met-tRNA(i)(Met)) pathway but required expression of euk

17 Met) ternary complex on base-pairing between Met-tRNA(i)(Met) and the start codon.

18 and eIF2 in eukaryotes are GTPases that bind Met-tRNA(i)(Met) to the small ribosomal subunit.

19 Translation initiation factor 2 (eIF2) binds Met-tRNA(i)(Met) to the 40S ribosomal subunit, and previ

20 In vitro, the MFC-GTP binds Met-tRNA(i) and delivers the tRNA to the ribosome at the

21 , a translation initiation factor that binds Met-tRNA(i), tryptic peptides from rabbit reticulocyte e

22 1:1 stoichiometry with respect to the bound Met-tRNA(i).

23 ynthetase (MRS) and production of a chimeric Met-tRNA(Ile) that would compromise translational fideli

24 IF3/eIF1 in a multifactor complex containing Met-tRNA(i)(Met).

25 F2 is a heterotrimer that binds and delivers Met-tRNA(i)(Met) to the 40 S ribosomal subunit in a GTP-

26 terotrimeric complex that binds and delivers Met-tRNA(i)(Met) to the 40 S ribosomal subunit in a GTP-

27 terial ribosomes, are critical for efficient Met-tRNA(i)(Met) binding and AUG selection in eukaryotes

28 In cooperation with eIF3, eIF1, and eIF1A, Met-tRNA(Met)(i)/eIF2/GTP binds to 40S subunits yielding

29 preinitiation complex (40 S.eIF3.eIF1.eIF1A.Met-tRNA(i).eIF2.GTP).

30 itiation factor (eIF) 3 and the ternary eIF2/Met-tRNA(i)(Met)/GTP complex and subsequently domain II

31 n eukaryotic translation initiation, eIF2GTP-Met-tRNA(i)(Met) ternary complex (TC) interacts with eIF

32 S preinitiation complex (40S.eIF1.eIF1A.eIF3.Met-tRNA(i).eIF2.GTP) and the subsequent binding of the

33 tRNA genes, the 5S rRNA gene (rrf), and an f-Met tRNA gene.

34 m of rrs and immediately downstream of the f-Met tRNA gene.

35 initiation was reduced with the fluorophore-Met-tRNA(f) compared with fMet-tRNA(f) with pyrene havin

36 s systems that can separate fMet-tRNA(fMet), Met-tRNA(fMet), and tRNA(fMet) shows that there is no fo

37 version of GTP to GDP on eIF2's affinity for Met-tRNA(i) in solution.

38 at-treated prt1-1 extracts are defective for Met-tRNA(i)Met binding to 40S subunits, and we also obse

39 Thus, three distinct pathways for Met-tRNA(i) delivery to the 40S ribosomal subunit are id

40 d in cell lysates, it may be responsible for Met-tRNA(i)-40S ribosome binding in vivo, possibly toget

41 romoting the GTP-dependent binding of formyl-Met-tRNA to the small subunit of either E. coli or bovin

42 spiratory defect in cells lacking formylated Met-tRNA(f)(Met) due to loss of the MIS1 gene that encod

43 factor might be unnecessary when formylated Met-tRNA(f)(Met) is present but becomes essential when o

44 requiring hydrolysis of GTP in the eIF2*GTP*Met-tRNA(i)(Met) ternary complex (TC) and subsequent P(i

45 lerate the rate of ternary complex (eIF2*GTP*Met-tRNA(i)(Met)) binding to 40S but only eIF1A stabiliz

46 en implicated in recruitment of the eIF2-GTP-Met-tRNA i Met ternary complex (TC) and ribosomal scanni

47 , which stimulates formation of the eIF2-GTP-Met-tRNA(i)(Met) ternary complex (TC) in a manner inhibi

48 nd indirectly through eIF5 with the eIF2-GTP-Met-tRNA(i)(Met) ternary complex (TC) to form the multif

49 of eIF1A impairs recruitment of the eIF2-GTP-Met-tRNA(i)(Met) ternary complex to 40S subunits, elimin

50 multifactor complex (eIF1-eIF3-eIF5-eIF2-GTP-Met-tRNA(i)(Met)).

51 ation complexes containing eIF3 and eIF2-GTP-Met-tRNA(iMet) to bind directly to the initiation codon,

52 e-initiator methionyl transfer RNA (eIF2.GTP.Met-tRNA(i )(Met)).

53 bly of the Saccharomyces cerevisiae eIF2.GTP.Met-tRNA(i) ternary complex and have determined the effe

54 GCN4 translation due to limiting of eIF2.GTP.Met-tRNA(i)(Met) ternary complex binding to the ribosome

55 primarily to regulate the level of eIF2.GTP.Met-tRNA(i)(Met) ternary complexes in vivo.

56 he eukaryotic initiation factor 2 (eIF2)/GTP/Met-tRNA(iMet) ternary complex (TC) and promotes scannin

57 aryotic translation initiation, the eIF2.GTP/Met-tRNA(i)(Met) ternary complex (TC) binds the eIF3/eIF

58 hroughout initiation: it stimulates eIF2/GTP/Met-tRNA(i)(Met) attachment to 40S ribosomal subunits, s

59 First, eIF5 binds eIF2/GTP/Met-tRNA(i)(Met) ternary complex (TC), promoting its rec

60 F5 stimulates GTP hydrolysis by the eIF2/GTP/Met-tRNA(i)(Met) ternary complex on base-pairing between

61 c initiation factor (eIF) 3 and the eIF2/GTP/Met-tRNA(i)(Met) ternary complex.

62 t interact with eIF1, eIF5, and the eIF2/GTP/Met-tRNA(i)(Met) ternary complex.

63 th enhance ribosomal recruitment of eIF2/GTP/Met-tRNA(i)(Met), but have opposite effects on the strin

64 irectly implicating eIF5-CTD in the eIF2/GTP/Met-tRNA(i)Met ternary complex binding process required

65 canonical UUG codon, presumably by impairing Met-tRNA(i)(Met) binding.

66 ma-N135D GTP-binding domain mutation impairs Met-tRNA(i)(Met) binding and causes a Sui(-) phenotype.

67 An eIF2gamma-A219T mutation impairs Met-tRNA(i)(Met) binding but unexpectedly enhances the f

68 to some extent utilize formylated initiator Met-tRNA to initiate protein synthesis and that initiati

69 ation in vivo of the mitochondrial initiator Met-tRNA in these strains.

70 eas a formylation-defective mutant initiator Met-tRNA, which binds to MTF with approximately the same

71 complex using in vitro binding of initiator Met-tRNA (as Met-tRNA(i).eIF2.GTP ternary complex) to 40

72 ng 80S ribosomes in the absence of initiator Met-tRNA(i) or any canonical initiation factors, from a

73 teraction between the anticodon of initiator Met-tRNA, associated with eIF2-GTP and 40S ribosomal sub

74 a tRNA-like IRES directs precise, initiator Met-tRNA-independent translation of two overlapping ORFs

75 t formation of 40S or 80S ribosome initiator Met-tRNA-AUG initiation complexes in vitro or on the pur

76 We show that the substrate initiator Met-tRNA protects MTF against trypsin cleavage, whereas

77 stem and the 3'-end region of the initiator Met-tRNA against cleavage by double and single strand-sp

78 In contrast, the initiator Met-tRNA is formylated in the respective "wild-type" par

79 eIF2 is a GTPase that delivers the initiator Met-tRNA to the P site on the small ribosomal subunit du

80 anticodon interactions between the initiator Met-tRNA(i)(Met) and the mRNA.

81 stly with the acceptor stem of the initiator Met-tRNA, which contains the critical determinants for f

82 mylation) are not protected by the initiator Met-tRNA.

83 rus can form 80S ribosomes without initiator Met-tRNA, eIF2, or GTP hydrolysis, with a CCU triplet in

84 lular eIF2 forms a complex with eIF5 lacking Met-tRNA(i)(Met), and here we investigate its physiologi

85 d both by eIF2/eIF3- and eIF5B/eIF3-mediated Met-tRNA(iMet) recruitment were destabilized by eIF1, di

86 Since an MFC-Met-tRNA(i) complex is detected in cell lysates, it may

87 portance of the formylation of mitochondrial Met-tRNA for the interaction with IF-2(mt) was investiga

88 icates that the formylation of mitochondrial Met-tRNA specifies its participation in initiation throu

89 will not effectively stimulate mitochondrial Met-tRNA binding to mitochondrial ribosomes, exhibiting

90 lated is not formylated by the mitochondrial Met-tRNA transformylase preventing its function in initi

91 dergoes an induced fit in the functional MTF.Met-tRNA complex but not in the nonfunctional one.

92 lso depends on formation of a functional MTF.Met-tRNA complex.

93 s cerevisiae can initiate with nonformylated Met-tRNA(f)(Met), as demonstrated in yeast mutants in wh

94 We could rescue 40S binding of Met- tRNA(i)Met and mRNA, and translation of luciferase

95 conformation to enable full accommodation of Met-tRNA(i)(Met) in the P site for AUG selection.

96 to the evolution of splicing.The binding of Met-tRNA to ribosomes is mediated by a GTP-binding prote

97 ut in vitro formylation increases binding of Met-tRNA(f)(Met) to translation initiation factor 2 (IF2

98 ctor complex in vivo, reduced the binding of Met-tRNA(i)(Met) and mRNA to 40S subunits in vitro.

99 propose that eIF5-CTD stimulates binding of Met-tRNA(i)(Met) and mRNA to 40S subunits through intera

100 n eIF2gamma subtly alter the conformation of Met-tRNA(i)(Met) on the 40S subunit and thereby affect t

101 The delivery of Met-tRNA(i) to the 40S ribosomal subunit is thought to o

102 iol and activated methionine, in the form of Met-tRNA, are incubated with MetRS.

103 ly in purine biosynthesis and formylation of Met-tRNA and indirectly in the biosynthesis of methionin

104 ty of start codon recognition independent of Met-tRNA(i)(Met) binding affinity.

105 A portion of Met-tRNA(Met) is formylated for initiation, whereas the

106 d SE2, was found to stimulate recruitment of Met-tRNA(i)(Met) in the ternary complex (TC) with eIF2.G

107 te efficient eIF2-independent recruitment of Met-tRNA(Met)(i) to 40S/mRNA complexes, if attachment of

108 ve interaction with the methionine moiety on Met-tRNA(i) that is disrupted when GTP is replaced with

109 , the NTT does not clash with either mRNA or Met-tRNA(i)(Met), consistent with its suggested role in

110 antly alter binding of guanine nucleotide or Met-tRNA(i)(Met) ligands by eIF2 in vitro.

111 ting a 50-fold preference for fMet-tRNA over Met-tRNA in this assay.

112 ic of mobile elements, including a potential Met-tRNA priming site, similar to that found in retrotra

113 report that eIF5B or eIF5B/eIF3 also promote Met-tRNA(iMet) binding to IRES-40S complexes, forming 48

114 ses two functions: synthetic, which provides Met-tRNA for protein synthesis, and editing, which rejec

115 n by eIF2 phosphorylation, despite requiring Met-tRNA(Met)(i).

116 A208V and A382V suppressor mutations restore Met-tRNA(i)(Met) binding affinity and cell growth; howev

117 f the ribosome, with initiator transfer RNA (Met-tRNA(i)(Met)) positioned over the start codon of mes

118 osin, pyrene, or coumarin attached to [(35)S]Met-tRNA(f).

119 anslational defect suggests eIF5B stabilizes Met-tRNA(i)(Met) binding and that GTP hydrolysis by eIF5

120 C) with eIF1, eIF2, and eIF5 that stimulates Met-tRNA(i)(Met) binding to 40S ribosomes and promotes s

121 er affinity for mitochondrial fMet-tRNA than Met-tRNA, using either the native mitochondrial tRNA(Met

122 40S ribosomes, creating the possibility that Met-tRNA(i) might bind directly to such 40S-factor compl

123 alysis to study the interactions between the Met-tRNA(fMet) and MTF in solution.

124 vitro or on the puromycin reactivity of the Met-tRNA in the 80S initiation complex.

125 MFC-GDP shows a greatly reduced affinity to Met-tRNA(i) compared to that for eIF2-GDP, suggesting th

126 mulates the GTPase activity of eIF2 bound to Met-tRNA(i)(Met), and its C-terminal domain (eIF5-CTD) b

127 nctions in binding methionyl initiator tRNA (Met-tRNA(i)(Met)) to the ribosome and in selecting AUG c

128 te it delivers the methionyl initiator tRNA (Met-tRNA(i)) to the small ribosomal subunit and releases

129 recruitment of aminoacylated initiator tRNA (Met-tRNA(Met)(i)) by eukaryotic initiation factor eIF2.

130 formylation of the initiator methionyl-tRNA (Met-tRNA(fMet)).

131 ect binding of the initiator methionyl-tRNA (Met-tRNA(i)) to 40 S ribosomal subunits in a codon-depen

132 N-Formylation of initiator methionyl-tRNA (Met-tRNA(Met)) by methionyl-tRNA formyltransferase (MTF)

133 fic formylation of initiator methionyl-tRNA (Met-tRNA) by methionyl-tRNA formyltransferase (MTF) is i

134 E. coli enzyme and initiator methionyl-tRNA (Met-tRNA) by using two complementary protection experime

135 ctor 2 (eIF2), the initiator methionyl-tRNA (Met-tRNA), and GTP is a critical step in translation ini

136 Compared to human tRNA gene promoters (tRNA(Met), tRNA(Val)), the human small nuclear RNA U6 gene (U

137 as the primer, can be forced to utilize tRNA(Met), tRNA(1,2)(Lys), tRNA(His), or tRNA(Glu), although

138 ons on the stability of methionylated tRNAi (Met-tRNA(i)) binding (in the ternary complex [TC] with e

139 The NTT and CTT both thread under Met-tRNA(i)(Met) reaching into the P-site.

140 ondrial IF2 (mIF2) in utilizing unformylated Met-tRNA(f)(Met).

141 e 40 S subunit in a multifactor complex with Met-tRNA(i)(Met), eIF2, eIF3, and eIF5 and binds near th

142 bility of eIF3 to stabilize the eIF2 x GTP x Met-tRNA(i) ternary complex.