June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
Crystal structure of Xenopus Arrestin 1 suggests multiple, competing dimer interfaces
Author Affiliations & Notes
  • Cassandra Barnes
    Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, New York, United States
  • David Salom
    Ophthalmology, Gavin Herbert Eye Institute, Irvine, California, United States
  • Michael Cosgrove
    Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, New York, United States
  • Philip David Kiser
    Physiology & Biophysics, University of California Irvine, Irvine, California, United States
    Ophthalmology, Gavin Herbert Eye Institute, Irvine, California, United States
  • Peter D Calvert
    Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, New York, United States
    Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, New York, United States
  • Footnotes
    Commercial Relationships   Cassandra Barnes None; David Salom None; Michael Cosgrove None; Philip Kiser None; Peter Calvert None
  • Footnotes
    Support  R01-EY018421, R01-EY028303
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 4491. doi:
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    • Get Citation

      Cassandra Barnes, David Salom, Michael Cosgrove, Philip David Kiser, Peter D Calvert; Crystal structure of Xenopus Arrestin 1 suggests multiple, competing dimer interfaces. Invest. Ophthalmol. Vis. Sci. 2023;64(8):4491.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Mammalian photoreceptor arrestin 1 (mArr1) self-associates into dimers and tetramers. We have shown that amphibian Arr1 oligomerization is limited to dimers. To identify structural differences from mArr1 that could explain the difference in the extent of oligomerization, we obtained crystal structures of Xenopus laevis Arr1 (xArr1). We employed site directed mutagenesis to validate the potential dimerization interfaces identified.

Methods : Recombinant WT xArr1 was expressed in E. coli, affinity purified and crystalized by vapor diffusion. Crystals were subjected to X-ray diffraction and xArr1 structures in two different space groups were solved at 2.5 and 2.9 Å. Residues at putative interaction interfaces were mutagenized and xArr1 mutants were subjected to analytical ultracentrifugation (AUC) and analytical size exclusion chromatography. AUC results were analyzed to determine oligomer formation and dissociation constants.

Results : Several potential dimer interfaces were identified in the two crystal forms. The dimer interface with the highest complexation significance score (CSS = 1) bears remarkable similarity to the C-C interface of the previously proposed mArr1 tetramer model [1], but with some key differences. First, the arrangement of the C-C dimer interface of xArr1 places one of the analogous phenylalanine residues (F193) required for mArr1 self-association outside of the interface; thus, it is not surprising that mutating this residue has no impact on xArr1 dimer formation. Second, the orientation of xArr1 protomers in the C-C dimer are slightly shifted and rotated as compared to mArr1. This difference may explain the inability of xArr1 to form tetramers. Intriguingly, a second possible C-N dimer interaction involves W190, a residue which appears to be important for the predicted C-C interface.

1. Hanson, S.M., et al., A model for the solution structure of the rod arrestin tetramer. Structure, 2008. 16(6): p 924-34.

Conclusions : Several potential dimer interfaces are predicted from the xArr1 crystal structures. Interestingly, one of these, which is remarkably similar to the C-C dimer predicted for the solution structure of mArr1, has never been obtained in mArr1 crystals. At least one other potential C-N dimer form was identified in the xArr1 crystals that may compete with the CC-dimer. Ongoing mutagenesis experiments will resolve which of these dimers form in solution.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

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