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Nikolai Artemyev, Liping Yu, Ravi Yadav; Role of the TPR-domain of AIPL1 in its function as PDE6 chaperone. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4932.
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AIPL1 is a specialized chaperone of phosphodiesterase-6 (PDE6) in rods and cones. Mutations in AIPL1 cause Leber congenital amaurosis type 4 (LCA4), a severe form of childhood blindness. The AIPL1-FKBP domain contributes to the chaperone activity by binding the C-terminal prenyl modifications of PDE6. Here, we have investigated the role of the AIPL1-TPR domain
X-ray crystallography was used to solve the crystal structure of the AIPL1-TPR domain. NMR spectroscopy was employed to make the backbone and sidechain assignments of the TPR domain of AIPL1 by using an uniformly 15N- and 13C-labeled AIPL1-TPR obtained from E. coli cell culture grown in a minimal medium containing 15NH4Cl and 13C6-glucose. Then, NMR chemical shift mapping technique was used to determine the PDE6 regulatory Pγ-subunit binding site on AIPL1-TPR. Fluorescence-binding assays and biolayer interferometry were used to localize the AIPL1-TPR binding site on the Pγ-subunit. Dynamic light scattering was also used to study thermal stability of AIPL1 in the presence of Pγ peptides
We identified a novel interaction between the AIPL1-TPR domain and the regulatory Pγ-subunit of PDE6. The AIPL1-TPR binding site of Pγ was localized to the C-terminal region of Pγ with the residues Pγ70-74 playing a critical role in the interaction. A working model of the complex of AIPL1-TPR with the Pγ C-terminus has been generated by mapping the chemical shift perturbations of AIPL1-TPR observed upon titrating Pγ onto the atomic structure of the TPR domain. Currently, this model is being probed with mutational analysis of the partner proteins
The interaction between the AIPL1-TPR domain and the C-terminal region of Pγ is likely to be critical for the chaperone function of AIPL1, since Pγ serves as a potent co-chaperone with AIPL1 in the folding and assembly of PDE6
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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