The structure of the human tRNALys3 anticodon bound to the HIV genome is stabilized by modified nucleosides and adjacent mismatch base pairs

Abstract

Replication of human immunodeficiency virus (HIV) requires base pairing of the reverse transcriptase primer, human tRNALys3, to the viral RNA. Although the major complementary base pairing occurs between the HIV primer binding sequence (PBS) and the tRNA's 3′-terminus, an important discriminatory, secondary contact occurs between the viral A-rich Loop I, 5′-adjacent to the PBS, and the modified, U-rich anticodon domain of tRNALys3. The importance of individual and combined anticodon modifications to the tRNA/HIV-1 Loop I RNA's interaction was determined. The thermal stabilities of variously modified tRNA anticodon region sequences bound to the Loop I of viral sub(sero)types G and B were analyzed and the structure of one duplex containing two modified nucleosides was determined using NMR spectroscopy and restrained molecular dynamics. The modifications 2-thiouridine, s2U34, and pseudouridine, Ψ39, appreciably stabilized the interaction of the anticodon region with the viral subtype G and B RNAs. The structure of the duplex results in two coaxially stacked A-form RNA stems separated by two mismatched base pairs, U162•Ψ39 and G163•A38, that maintained a reasonable A-form helix diameter. The tRNA's s2U34 stabilized the interaction between the A-rich HIV Loop I sequence and the U-rich anticodon, whereas the tRNA's Ψ39 stabilized the adjacent mismatched pairs.

A novel method of liquid chromatography–tandem mass spectrometry combined with chemical derivatization for the determination of ribonucleosides in urine

“…The excretion of ribonucleosides is not affected by age and diet [2]. But elevated levels of ribonucleosides were observed in patients with physical illness, such as urogenital cancer [3,4], hepatocellular carcinoma [5,6], breast cancer [7], acquired immunodeficiency diseases (AIDS) [8,9] and severe combined immunodeficiency diseases (SCID) [10]. Therefore, the levels of urinary ribonucleosides can be considered as a signal of disease status, especially as potential biomarkers for cancer diagnosis.…”

Thermodynamic contribution and nearest-neighbor parameters of pseudouridine-adenosine base pairs in oligoribonucleotides

“…This is thought to be due to stabilization imparted by Ψ's novel hydrogen bonding capabilities, resulting in more accurate and stable recognition interactions between tRNA and the ribosome (Davis and Poulter 1991). Notably, this property is exploited by the HIV genome, which relies on the stabilization afforded by the pseudouridylation of tRNA Lys3 to act as a reverse transcriptase primer (Bilbille et al 2009). Ψ may also play a prominent role in spliceosomal RNA responsible for gene regulation.…”

Modifications Modulate Anticodon Loop Dynamics and Codon Recognition of E. coli tRNAArg1,2

“…35 The sulfur atom at position 2 of pyrimidines is known to stabilize stacking interactions by its greater polarizability than oxygen and to promote the C3′-endo, anti-conformation of the nucleoside in RNA. 36 Since the bases of nucleosides 31, 32 and 33 should be stacked in the classic U-turn conformation, one wonders how an increase in stacking induced by the 2-thiocytidine may influence the base pairing of C 32 with A 38 . The structure of the E. coli tRNA Arg1,2 ICG -s 2 C 32 free in solution may be influenced by the s 2 C 32 modification because a change from C 32 to s 2 C 32 results in an altered accessibility of the anticodon loop to nuclease S1 to a pattern more indicative of an initiator than an elongator tRNA.…”