Adaptation of the Romanomermis culicivorax CCA-Adding Enzyme to Miniaturized Armless tRNA Substrates.

The mitochondrial genome of the nematode Romanomermis culicivorax encodes for miniaturized hairpin-like tRNA molecules that lack D- as well as T-arms, strongly deviating from the consensus cloverleaf. The single tRNA nucleotidyltransferase of this organism is fully active on armless tRNAs, while the human counterpart is not able to add a complete CCA-end. Transplanting single regions of the Romanomermis enzyme into the human counterpart, we identified a beta-turn element of the catalytic core that-when inserted into the human enzyme-confers full CCA-adding activity on armless tRNAs. This region, originally identified to position the 3'-end of the tRNA primer in the catalytic core, dramatically increases the enzyme's substrate affinity. While conventional tRNA substrates bind to the enzyme by interactions with the T-arm, this is not possible in the case of armless tRNAs, and the strong contribution of the beta-turn compensates for an otherwise too weak interaction required for the addition of a complete CCA-terminus. This compensation demonstrates the remarkable evolutionary plasticity of the catalytic core elements of this enzyme to adapt to unconventional tRNA substrates.

[Electrophoretic analysis of isoenzyme and protein patterns of Romanomermis yunanensis and R. culicivorax (Nematoda:Mermithidae)].

Isoenzyme electrophoretic patterns of lactate dehydrogenase (LDH), malate dehydrogenase (MDH), esterase (EST) and peroxidase (PO) of post-parasitic juveniles and adults of Romanomermis yunanensis and R. culicivorax were analysed by disc electrophoresis, respectively. The protein patterns of the two species of Romanomermis were compared by vertical slab SDS polyacrylamide gel electrophoresis. The results showed marked differences in isoenzyme patterns (number of bands and/or Rf) of LDH and MDH between the two species of Romanomermis, but the similar positions of isoenzyme bands denoted a certain genetic relationship between them. No band was found in the isoenzyme patterns of PO and EST. Distinct differences in protein pattern were observed between the two species of Romanomermis, but some common bands in pattern reflected the phylogenetic relationships of these species. The differences of isoenzyme and protein patterns observed in this study have provided the reason for identifying and differentiating the two species of Romanomermis at the molecular level.

Sequence amplification and gene rearrangement in parasitic nematode mitochondrial DNA.

The nematode Romanomermis culicivorax, an obligate mosquito parasite, possesses a 26 kilobase (kb) mitochondrial genome. The unusually large size is due to transcriptionally active DNA sequences present as 3.0 kb direct tandem repeats and as inverted portions of the repeating unit located elsewhere in the mitochondrial DNA (mtDNA). The genome rearrangements involved in establishing this unusual sequence organization may have dramatically altered conventional mitochondrial gene order. Genes for subunits of the cytochrome c oxidase complex (COI and COII) are normally closely linked in animal mtDNAs, but are separated by approximately 8 kb in this mitochondrial genome.