BACKGROUND: Metazoan mitochondrial (mt) tRNAs are structurally quite different from the canonical cloverleaf secondary structure. The mammalian mt tRNASerGCU for AGY codons (Y = C or U) lacks the entire D arm, whereas tRNASerUGA for UCN codons (N = A, G, C or U) has an extended anti-codon stem. It has been a long-standing problem to prove experimentally how these tRNAsSer work in the mt translation system. RESULTS: To solve the above-mentioned problem, we examined their translational abilities in an in vitro bovine mitochondrial translation system using transcripts of altered tRNASer analogues derived from bovine mitochondria. Both tRNASer analogues had almost the same ability to form ternary complexes with mt EF-Tu and GTP. The D-arm-lacking tRNASer GCU analogue had considerably lower translational activity than the tRNASerUGA analogue and produced mostly short oligopeptides, up to a tetramer. In addition, tRNASerGCU analogue was disfavoured by the ribosome when other tRNAs capable of decoding the cognate codon were available. CONCLUSION: Both mt tRNASerGCU and tRNASerUGA analogues with unusual secondary structure were found to be capable of translation on the ribosome. However, the tRNASerGCU analogue has some molecular disadvantage on the ribosome, which probably derives from the lack of a D arm.

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