Nucleic-acid-binding properties of the C2-L1Tc nucleic acid chaperone encoded by L1Tc retrotransposon.
It has been reported previously that the C2-L1Tc protein located in the Trypanosoma cruzi LINE (long interspersed nuclear element) L1Tc 3' terminal end has NAC (nucleic acid chaperone) activity, an essential activity for retrotransposition of LINE-1. The C2-L1Tc protein contains two cysteine motifs of a C2H2 type, similar to those present in TFIIIA (transcription factor IIIA). The cysteine motifs are flanked by positively charged amino acid regions. The results of the present study show that the C2-L1Tc recombinant protein has at least a 16-fold higher affinity for single-stranded than for double-stranded nucleic acids, and that it exhibits a clear preference for RNA binding over DNA. The C2-L1Tc binding profile (to RNA and DNA) corresponds to a non-co-operative-binding model. The zinc fingers present in C2-L1Tc have a different binding affinity to nucleic acid molecules and also different NAC activity. The RRR and RRRKEK [NLS (nuclear localization sequence)] sequences, as well as the C2H2 zinc finger located immediately downstream of these basic stretches are the main motifs responsible for the strong affinity of C2-L1Tc to RNA. These domains also contribute to bind single- and double-stranded DNA and have a duplex-stabilizing effect. However, the peptide containing the zinc finger situated towards the C-terminal end of C2-L1Tc protein has a slight destabilization effect on a mismatched DNA duplex and shows a strong preference for single-stranded nucleic acids, such as C2-L1Tc. These results provide further insight into the essential properties of the C2-L1Tc protein as a NAC.
Heras
Sara R
SR
Departamento de Biología Molecular, Instituto de Parasitología y Biomedicina López Neyra, CSIC, 18001 Granada, Spain.
Thomas
M Carmen
MC
Macias
Francisco
F
Patarroyo
Manuel E
ME
Alonso
Carlos
C
López
Manuel C
MC
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D004247:DNA / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D007700:Kinetics; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009696:Nucleic Acids / Q000378:metabolism; D011485:Protein Binding; D012313:RNA / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D018626:Retroelements; D014349:Trypanosoma cruzi / Q000235:genetics; D016335:Zinc Fingers / Q000235:genetics
D004277:DNA, Single-Stranded; D018832:Molecular Chaperones; D009696:Nucleic Acids; D011994:Recombinant Proteins; D018626:Retroelements; D012313:RNA; D004247:DNA