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The structure of Sky1p reveals a novel mechanism for constitutive activity

Nature Structural Biology volume 8, pages 176–183 (2001)Cite this article

Abstract

Sky1p is the only member of the SR protein kinase (SRPK) family in Saccharomyces cerevisiae. SRPKs are constitutively active kinases that display remarkable substrate specificity and have been implicated in RNA processing. Here we present the three-dimensional structure of a fully active truncated Sky1p. Analysis of the structure and structure-based functional studies reveal that the C-terminal tail, an unusual Glu residue located in the P+1 loop, and a unique mechanism for the positioning of helix αC act together to render Sky1p constitutively active. We have modeled a substrate peptide bound to Sky1p. The modeled complex combined with mutagenesis studies illustrate the molecular basis for substrate recognition by this kinase and suggest a mechanism by which SRPKs catalyze a sequential phosphorylation reaction of the consecutive RS dipeptide repeats characteristic of mammalian SRPK substrates.

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Figure 1: Sequence alignment and secondary structure of an active Sky1p construct.
Figure 2: Overall structure of Sky1p.
Figure 3: The active conformation of Sky1p and the orientation of helix αC.
Figure 4: Activation loop conformation and the C-terminal tail.
Figure 5: The role of Gln 566 in maintaining the active conformation.
Figure 6: Substrate specificity of Sky1p.

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Acknowledgements

We wish to thank T. Huxford and the other members of the Ghosh lab for comments on the manuscript and N. Nguyen for his technical assistance at the UCSD X-ray source. B.N. is supported by a fellowship from the Molecular Biophysics Training Program. C.Y. is a National Cancer Institute graduate trainee for Cancer Cell Biology. X.-D.F. is a Leukemia and Lymphoma Scholar; G.G. is an Alfred P. Sloan fellow.

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Authors and Affiliations

  1. Department of Chemistry and Biochemistry, University of California, San Diego, 9500 Gilman Drive, La Jolla, 92130, California, USA

    Brad Nolen, J. Andrew McCammon & Gourisankar Ghosh

  2. Department of Cellular and Molecular Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, 92130, California, USA

    Chi Y. Yun & Xiang-Dong Fu

  3. Department of Pharmacology and Howard Hughes Medical Institute, University of California, San Diego, 9500 Gilman Drive, La Jolla, 92130, California, USA

    Chung F. Wong & J. Andrew McCammon

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Correspondence to Gourisankar Ghosh.

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Nolen, B., Yun, C., Wong, C. et al. The structure of Sky1p reveals a novel mechanism for constitutive activity. Nat Struct Mol Biol 8, 176–183 (2001). https://doi.org/10.1038/84178

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