Structure of the two most C-terminal RNA recognition motifs of PTB using segmental isotope labeling.
The polypyrimidine tract binding protein (PTB) is a 58 kDa protein involved in many aspects of RNA metabolism. In this study, we focused our attention on the structure of the two C-terminal RNA recognition motifs (RRM3 and RRM4) of PTB. In a previous study, it was found that the two RRMs are independent in the free state. We recently determined the structure of the same fragment in complex with RNA and found that the two RRMs interact extensively. This difference made us re-evaluate in detail the free protein structure and in particular the interdomain interface. We used a combination of NMR spectroscopy and segmental isotopic labeling to unambiguously study and characterize the interdomain interactions. An improved segmental isotopic labeling protocol was used, enabling us to unambiguously identify 130 interdomain NOEs between the two RRMs and to calculate a very precise structure. The structure reveals a large interdomain interface, resulting in a very unusual positioning of the two RRM domains relative to one another.
Vitali
Francesca
F
Institute for Molecular Biology and Biophysics, Swiss Federal Institute of Technology Zurich, ETH-Hönggerberg, Zürich, Switzerland.
Henning
Anke
A
Oberstrass
Florian C
FC
Hargous
Yann
Y
Auweter
Sigrid D
SD
Erat
Michèle
M
Allain
Frédéric H-T
FH
D020816:Amino Acid Motifs / Q000235:genetics; D002247:Carbon Isotopes; D034441:Heterogeneous-Nuclear Ribonucleoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007553:Isotope Labeling; D009682:Magnetic Resonance Spectroscopy; D016296:Mutagenesis; D009587:Nitrogen Isotopes; D038941:Polypyrimidine Tract-Binding Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism
D002247:Carbon Isotopes; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D009587:Nitrogen Isotopes; C450006:PTBP1 protein, human; D012333:RNA, Messenger; D038941:Polypyrimidine Tract-Binding Protein