Trypanosoma brucei gBP21. An arginine-rich mitochondrial protein that binds to guide RNA with high affinity.
RNA editing in Trypanosoma brucei is a mitochondrial RNA processing reaction that results in the insertion and deletion of uridylate residues into otherwise untranslatable mRNAs. The process is directed by guide RNAs which function to specify the edited sequence. RNA editing in vitro requires mitochondrial protein extracts and guide RNAs have been identified as part of high molecular weight ribonucleoprotein complexes. Within the complexes, the RNAs are in close contact with several mitochondrial proteins and here we describe the isolation and cloning of a gRNA-interacting polypeptide from Trypanosoma brucei. The protein was named gBP21 for guide RNA-binding protein of 21 kDa. gBP21 shows no homology to proteins in other organisms, it is arginine-rich and binds to gRNA molecules with a dissociation constant in the nanomolar range. The protein does not discriminate for differences in the primary structures of gRNAs and thus likely binds to higher order structural features common to all gRNA molecules. gBP21 binding does not perturb the overall structure of gRNAs but the gRNA/gBP21 ribonucleoprotein complex is more stable than naked guide RNAs. Although the protein is arginine-rich, the free amino acid or an arginine-rich peptide were not able to inhibit the association to the RNAs. In contrast, the gRNA-gBP21 complex formation was sensitive to potassium and ammonium cations, thus indicating a contribution of ionic contacts to the binding.
Köller
J
J
Laboratorium für molekulare Biologie, Genzentrum der LMU München am MPI für Biochemie, 82152 Martinsried, Germany.
Müller
U F
UF
Schmid
B
B
Missel
A
A
Kruft
V
V
Stuart
K
K
Göringer
H U
HU
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000032:analysis*; D001483:Base Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011487:Protein Conformation; D015800:Protozoan Proteins; D017394:RNA, Guide, Kinetoplastida / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D014346:Trypanosoma brucei brucei / Q000737:chemistry*
D015800:Protozoan Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; C104816:gBP21 protein, Trypanosoma brucei; D001120:Arginine; D017394:RNA, Guide, Kinetoplastida