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Structure-function studies of FMRP RGG peptide recognition of an RNA duplex-quadruplex junction

Nature Structural & Molecular Biology volume 18, pages 796–804 (2011)Cite this article

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Abstract

We have determined the solution structure of the complex between an arginine-glycine-rich RGG peptide from the human fragile X mental retardation protein (FMRP) and an in vitro–selected guanine-rich (G-rich) sc1 RNA. The bound RNA forms a newly discovered G-quadruplex separated from the flanking duplex stem by a mixed junctional tetrad. The RGG peptide is positioned along the major groove of the RNA duplex, with the G-quadruplex forcing a sharp turn of R10GGGGR15 at the duplex-quadruplex junction. Arg10 and Arg15 form cross-strand specificity–determining intermolecular hydrogen bonds with the major-groove edges of guanines of adjacent Watson-Crick G•C pairs. Filter-binding assays on RNA and peptide mutations identify and validate contributions of peptide-RNA intermolecular contacts and shape complementarity to molecular recognition. These findings on FMRP RGG domain recognition by a combination of G-quadruplex and surrounding RNA sequences have implications for the recognition of other genomic G-rich RNAs.

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Figure 1: Sequence and NMR spectra of sc1 RNA and RGG peptide.
Figure 2: RNA resonance assignments in the RGG peptide–sc1 RNA complex.
Figure 3: Identification of G-tetrad alignments and assignment of through-bond correlations in the RGG peptide–sc1 RNA complex.
Figure 4: Solution structure of the RGG peptide–sc1 RNA complex and the architecture of the G-quadruplex and duplex-quadruplex junction.
Figure 5: Architecture of the G-quadruplex and duplex-quadruplex junction.
Figure 6: Details of intermolecular peptide-RNA interactions in the solution structure of the RGG peptide–sc1 RNA complex.
Figure 7: Assessment of the molecular determinants of the peptide and of the RNA for the FMRP RGG peptide–sc1 RNA interaction by filter-binding assay.

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Acknowledgements

We thank Y. Gosser and S. Pitt (Memorial Sloan-Kettering Cancer Center) for their participation at the early stage of the project, O. Mostovetsky, E.F. Stone and K.Y.S. Hung (Rockefeller University) for technical assistance, and A. Lash (Memorial Sloan-Kettering Cancer Center) for bioinformatics of the RG4R motif in the human genome. This research was supported by US National Institutes of Health grants CA049982 (to D.J.P.) and R01 HD040647 (to J.C.D.) and Singapore Biomedical Research Council grant 07/1/22/19/542 to A.T.P. D.J.P. is a member of the New York Structural Biology Center, supported by US National Institutes of Health grant GM66354.

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  1. Anh Tuân Phan, Vitaly Kuryavyi and Jennifer C Darnell: These authors contributed equally to this work.

Authors and Affiliations

  1. Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York, USA

    Anh Tuân Phan, Vitaly Kuryavyi, Alexander Serganov, Serge Ilin, Tanya Raslin, Anna Polonskaia & Dinshaw J Patel

  2. School of Physical and Mathematical Sciences, Nanyang Technological University, Singapore

    Anh Tuân Phan

  3. Laboratory of Molecular Neuro-Oncology, The Rockefeller University, New York, New York, USA

    Jennifer C Darnell, Cynthia Chen, David Clain & Robert B Darnell

  4. Howard Hughes Medical Institute, The Rockefeller University, New York, New York, USA

    Robert B Darnell

  5. Biomolecular NMR Center, Johns Hopkins University, Baltimore, Maryland, USA

    Ananya Majumdar

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Contributions

A.T.P., A.M. and S.I. were responsible for NMR studies, V.K. undertook the computations, and A.S., T.R. and A.P. prepared labeled NMR samples, all under the supervision of D.J.P. C.C., D.C. and J.C.D. did the filter-binding assays under the joint supervision of J.C.D. and R.B.D. The paper was written jointly by D.J.P., A.T.P., V.K., J.C.D. and R.B.D.

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Correspondence to Anh Tuân Phan, Jennifer C Darnell or Dinshaw J Patel.

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Phan, A., Kuryavyi, V., Darnell, J. et al. Structure-function studies of FMRP RGG peptide recognition of an RNA duplex-quadruplex junction. Nat Struct Mol Biol 18, 796–804 (2011). https://doi.org/10.1038/nsmb.2064

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