A 3-DIMENSIONAL WORKING MODEL FOR A GUIDE RNA FROM TRYPANOSOMA-BRUCEI

RNA editing in protozoan parasites is a mitochondrial RNA processing r eaction in which exclusively uridylate residues are inserted into, and less frequently deleted from, pre-mRNAs. Molecules central to the pro cess are so-called guide RNAs (gRNAs) which function as templates in t he reaction. For a detailed molecular understanding of the mechanism o f the editing process knowledge of structural features of gRNAs will b e essential. Here we report on a computer-assisted molecular modelling approach to construct the first three-dimensional gRNA model for gND7 -506, a ND7-specific gRNA from Trypanosoma brucei. The modelling proce ss relied on chemical modification and enzymatic probing data and was validated by in vitro mutagenesis experiments. The model predicts a re asonably compact structure, where two stem/loop secondary structure el ements are brought into close proximity by a triple A tertiary interac tion, forming a core element within the centre of the molecule. The mo del further suggests that the surface of the gRNA is primarily made up of the sugar-phoshate backbone. On the basis of the model, footprinti ng experiments of gND7-506 in a complex with the gRNA binding protein gBP21 could successfully be interpreted and provide a first picture fo r the assembly of gRNAs within a ribonucleoprotein complex.