Unique secondary and tertiary structural features of the eucaryotic selenocysteine tRNASec

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Abstract
Cotranslatlonal insertion of selenocysteine into selenoenzymes is mediated by a specialized transfer RNA, the tRNASec. We have carried out the determination of the solution structure of the eucaryotlc tRNASec. Based on the enzymatic and chemical probing approach, we show that the secondary structure bears a few unprecedented features like a 9 bp amlnoacld-, a 4 bp thymine- and a 6 bp dlhydrouridine-stems. Surprisingly, the eighth nucleotide, although being a uridine, Is base-paired and cannot therefore correspond to the single-stranded invariant U8 found in all tRNAs. Rather, experimental evidence led us to propose that the role of the invariant U8 is actually played by the tenth nucleotide which is an A, numbered A8 to Indicate this fact. The experimental data therefore demonstrate that the cloverleaf structure we derived experimentally resembles the hand-folded model proposed by Böck et al (ref. 3). Using the solution data and computer modelling, we derived a three-dimensional structure model which shows some unique aspects. Basically, A8, A14, U21 form a novel type of tertiary interaction in which A8 interacts with the Hoogsteen sites of A14 which itself forms a Watson-Crick pair with U21. No coherent model containing the canonical 15–48 interaction could be derived. Thus, the number of tertiary interactions appears to be limited, leading to an uncoupling of the variable stem from the rest of the molecule.