3T1Y: Structure Of The Thermus Thermophilus 30s Ribosomal Subunit Complexed With A Human Anti-Codon Stem Loop (Hasl) Of Transfer Rna Lysine 3 (Trnalys3) Bound To An Mrna With An Aag-Codon In The A-Site And Paromomycin

Citation:
Abstract
Human tRNA(Lys3)(UUU) (htRNA(Lys3)(UUU)) decodes the lysine codons AAA and AAG during translation and also plays a crucial role as the primer for HIV-1 (human immunodeficiency virus type 1) reverse transcription. The posttranscriptional modifications 5-methoxycarbonylmethyl-2-thiouridine (mcm(5)s(2)U(34)), 2-methylthio-N(6)-threonylcarbamoyladenosine (ms(2)t(6)A(37)), and pseudouridine (Psi(39)) in the tRNA's anticodon domain are critical for ribosomal binding and HIV-1 reverse transcription. To understand the importance of modified nucleoside contributions, we determined the structure and function of this tRNA's anticodon stem and loop (ASL) domain with these modifications at positions 34, 37, and 39, respectively (hASL(Lys3)(UUU)-mcm(5)s(2)U(34);ms(2)t(6)A(37);Psi(39)). Ribosome binding assays in vitro revealed that the hASL(Lys3)(UUU)-mcm(5)s(2)U(34);ms(2)t(6)A(37);Psi(39) bound AAA and AAG codons, whereas binding of the unmodified ASL(Lys3)(UUU) was barely detectable. The UV hyperchromicity, the circular dichroism, and the structural analyses indicated that Psi(39) enhanced the thermodynamic stability of the ASL through base stacking while ms(2)t(6)A(37) restrained the anticodon to adopt an open loop conformation that is required for ribosomal binding. The NMR-restrained molecular-dynamics-derived solution structure revealed that the modifications provided an open, ordered loop for codon binding. The crystal structures of the hASL(Lys3)(UUU)-mcm(5)s(2)U(34);ms(2)t(6)A(37);Psi(39) bound to the 30S ribosomal subunit with each codon in the A site showed that the modified nucleotides mcm(5)s(2)U(34) and ms(2)t(6)A(37) participate in the stability of the anticodon-codon interaction. Importantly, the mcm(5)s(2)U(34).G(3) wobble base pair is in the Watson-Crick geometry, requiring unusual hydrogen bonding to G in which mcm(5)s(2)U(34) must shift from the keto to the enol form. The results unambiguously demonstrate that modifications pre-structure the anticodon as a key prerequisite for efficient and accurate recognition of cognate and wobble codons.
PDB ID: 3T1YDownload
MMDB ID: 96706
PDB Deposition Date: 2011/7/22
Updated in MMDB: 2012/09
Experimental Method:
x-ray diffraction
Resolution: 2.8  Å
Source Organism:
Thermus thermophilus HB8
Similar Structures:
Biological Unit for 3T1Y: 23-meric; determined by author
Molecular Components in 3T1Y
Label Count Molecule
Proteins (20 molecules)
1
30S Ribosomal Protein S2(Gene symbol: rpsB)
Molecule annotation
1
30S Ribosomal Protein S3(Gene symbol: rpsC)
Molecule annotation
1
30S Ribosomal Protein S4(Gene symbol: rpsD)
Molecule annotation
1
30S Ribosomal Protein S5(Gene symbol: rpsE)
Molecule annotation
1
30S Ribosomal Protein S6(Gene symbol: rpsF)
Molecule annotation
1
30S Ribosomal Protein S7(Gene symbol: TTHA1696)
Molecule annotation
1
30S Ribosomal Protein S8
Molecule annotation
1
30S Ribosomal Protein S9
Molecule annotation
1
30S Ribosomal Protein S10(Gene symbol: rpsJ)
Molecule annotation
1
30S Ribosomal Protein S11(Gene symbol: TTHA1666)
Molecule annotation
1
30S Ribosomal Protein S12(Gene symbol: rpsL)
Molecule annotation
1
30S Ribosomal Protein S13(Gene symbol: rpsM)
Molecule annotation
1
30S Ribosomal Protein S14 Type Z
Molecule annotation
1
30S Ribosomal Protein S15(Gene symbol: rpsO)
Molecule annotation
1
30S Ribosomal Protein S16(Gene symbol: rpsP)
Molecule annotation
1
30S Ribosomal Protein S17
Molecule annotation
1
30S Ribosomal Protein S18
Molecule annotation
1
30S Ribosomal Protein S19(Gene symbol: rpsS)
Molecule annotation
1
30S Ribosomal Protein S20
Molecule annotation
1
30S Ribosomal Protein THX(Gene symbol: TTHA1396)
Molecule annotation
Nucleotides(3 molecules)
1
16S rRNA
Molecule annotation
1
mRNA A-site Fragment
Molecule annotation
1
tRNA ASL Human Lys3
Molecule annotation
Chemicals (189 molecules)
1
186
2
2
3
1
* Click molecule labels to explore molecular sequence information.

Citing MMDB
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