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Category: Microbial Genetics and Molecular Biology
The tRNA Identity Problem: Past, Present, and Future, Page 1 of 2
< Previous page | Next page > /docserver/preview/fulltext/10.1128/9781555818333/9781555810733_Chap16-1.gif /docserver/preview/fulltext/10.1128/9781555818333/9781555810733_Chap16-2.gifAbstract:
This chapter describes the current understanding of the structural features in tRNA that determine the specificity of the interaction with the aminoacyl-tRNA synthetase (aaRS), and outlines future research in this area. Early methods of sequence comparison to predict tRNA identity determinants relied only on the structural similarities among isoacceptor tRNAs discerned by visual inspection of the sequences. Positions where the same nucleotide occurs in all isoacceptor species were considered more predictive of tRNA identity than were positions where the nucleotide differed among the isoacceptors. The newer approach relies on computer analysis of tRNA sequences and identifies not only conserved nucleotide positions within a tRNA acceptor group, but also positions with more than one nucleotide that differ from the corresponding nucleotide positions in other tRNA acceptor groups. In addition, because the between-group variation is considered, the newer method provides information for single tRNA sequences and can perform impressively when only two tRNA isoacceptor sequences are known. The chapter summarizes experiments that define specificity determinants in tRNAs for several amino acids, and includes illustrations of computer predictions. The implications of the study results are discussed, and the chapter closes with an outline of future prospects.
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General tRNA structures. (A) Cloverleaf structure with bases replaced by numbers indicating the standard 76 nucleotide positions. The constant nucleotides are noted. The • symbol indicates Watson-Crick base pairings, and the thin line, other base pairings. B: L-shaped structure with shading of bases 16, 17, 20, 59, and 60 of the variable pocket (VP). Adapted from references 38 and 46 , with permission.
General tRNA structures. (A) Cloverleaf structure with bases replaced by numbers indicating the standard 76 nucleotide positions. The constant nucleotides are noted. The • symbol indicates Watson-Crick base pairings, and the thin line, other base pairings. B: L-shaped structure with shading of bases 16, 17, 20, 59, and 60 of the variable pocket (VP). Adapted from references 38 and 46 , with permission.
Histogram showing predicted determinants of tRNA identity obtained by analysis of 67 E. coli tRNA sequences. Histograms are shown for tRNAGly, which has five isoacceptors (A); tRNAAla, which has two isoacceptors (B); and tRNAPhe, which has one acceptor (C). Frequency of correlation on the vertical axis is a function of tRNA position on the horizontal axis. Cloverleaf parts are indicated on the horizontal axis. A = acceptor stem; D = D stem; C = anticodon stem; X = anticodon; and T = T stem. The middle anticodon nucleotide, residue 35, is indicated. The base and position number of an established specificity determinant is indicated above each data bar. Adapted from references 38 , 40 , and 44 , with permission.
Histogram showing predicted determinants of tRNA identity obtained by analysis of 67 E. coli tRNA sequences. Histograms are shown for tRNAGly, which has five isoacceptors (A); tRNAAla, which has two isoacceptors (B); and tRNAPhe, which has one acceptor (C). Frequency of correlation on the vertical axis is a function of tRNA position on the horizontal axis. Cloverleaf parts are indicated on the horizontal axis. A = acceptor stem; D = D stem; C = anticodon stem; X = anticodon; and T = T stem. The middle anticodon nucleotide, residue 35, is indicated. The base and position number of an established specificity determinant is indicated above each data bar. Adapted from references 38 , 40 , and 44 , with permission.
Cloverleaf arrangement of nucleotide sequences of E. coli tRNAs corresponding to amber suppressors, indicating standard position numbers for Gly (A), Phe (B), and Lys (C). Nucleotide modifications are not indicated. The ′ symbol represents an alignment gap. Adapted from reference 44 , with permission.
Cloverleaf arrangement of nucleotide sequences of E. coli tRNAs corresponding to amber suppressors, indicating standard position numbers for Gly (A), Phe (B), and Lys (C). Nucleotide modifications are not indicated. The ′ symbol represents an alignment gap. Adapted from reference 44 , with permission.
Comparison of composite nucleotide sequences of amber suppressors representing tRNAGly-like U73 mutants of tRNAPhe and tRNAArg and isoacceptors of tRNAGly (A); non-tRNAGly-like U73 mutants of tRNALys and tRNAGln (C); and nucleotides common to A and C (B). The positions where the two composites do not contain a common nucleotide are underlined and marked with the * symbol in (B). Incompletely specified nucleotides are designated as M = AC; R = AG; W = AU; S = CG; Y = CU; K = GU; V = ACG; H = ACU; D = AGU; B = CGU; and N = ACGU. An alignment gap is indicated by the ′ symbol. Lowercase letters indicate the position contains either an alignment gap or the indicated nucleotide. Adapted from reference 44 , with permission.
Comparison of composite nucleotide sequences of amber suppressors representing tRNAGly-like U73 mutants of tRNAPhe and tRNAArg and isoacceptors of tRNAGly (A); non-tRNAGly-like U73 mutants of tRNALys and tRNAGln (C); and nucleotides common to A and C (B). The positions where the two composites do not contain a common nucleotide are underlined and marked with the * symbol in (B). Incompletely specified nucleotides are designated as M = AC; R = AG; W = AU; S = CG; Y = CU; K = GU; V = ACG; H = ACU; D = AGU; B = CGU; and N = ACGU. An alignment gap is indicated by the ′ symbol. Lowercase letters indicate the position contains either an alignment gap or the indicated nucleotide. Adapted from reference 44 , with permission.
Histogram showing percentage of Ala inserted in suppressed protein by mutants of amber-suppressor tRNAAla. Panel A shows wild-type G3•U70 and the 15 possible base 3-base 70 mutant combinations. Panel B shows wild-type G3•U70, several mutants with a G•U wobble pair at position 2•71 or position 4•69, and a mutant with G3•U70 flanked by mutant base pairs. Reproduced from reference 39 , with permission.
Histogram showing percentage of Ala inserted in suppressed protein by mutants of amber-suppressor tRNAAla. Panel A shows wild-type G3•U70 and the 15 possible base 3-base 70 mutant combinations. Panel B shows wild-type G3•U70, several mutants with a G•U wobble pair at position 2•71 or position 4•69, and a mutant with G3•U70 flanked by mutant base pairs. Reproduced from reference 39 , with permission.
Model of variable pocket of yeast tRNAPhe. The acceptor stem and T stem and part of the D loop are shown as viewed by looking down from a point behind and to the right of the acceptor stem, as shown in Fig. 1B . Bases are represented as hatched circles, phosphates and sugars are shown as open circles, and hydrogen bonds are shown as dashed lines. Open arrows indicate points where the polynucleotide chain continues to other parts of the molecule. Note that the variable pocket is segregated from the constant nucleotides 18, 19, 54, 55, 56, and 58. Reproduced from reference 36 , with permission.
Model of variable pocket of yeast tRNAPhe. The acceptor stem and T stem and part of the D loop are shown as viewed by looking down from a point behind and to the right of the acceptor stem, as shown in Fig. 1B . Bases are represented as hatched circles, phosphates and sugars are shown as open circles, and hydrogen bonds are shown as dashed lines. Open arrows indicate points where the polynucleotide chain continues to other parts of the molecule. Note that the variable pocket is segregated from the constant nucleotides 18, 19, 54, 55, 56, and 58. Reproduced from reference 36 , with permission.
Structure of small RNAs, including amber- suppressor tRNAPhe (A), deleted derivative lacking the D region (B), and minihelix RNA (C). Modified nucleotides are not indicated. The ′ symbol indicates a deleted nucleotide. Adapted from reference 45 , with permission.
Structure of small RNAs, including amber- suppressor tRNAPhe (A), deleted derivative lacking the D region (B), and minihelix RNA (C). Modified nucleotides are not indicated. The ′ symbol indicates a deleted nucleotide. Adapted from reference 45 , with permission.