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Ravi Prakash Yadav, Anurima Majumder, Lokesh Gakhar, Nikolai Artemyev; Probing the role of the C-terminal region of human AIPL1. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4656.
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© ARVO (1962-2015); The Authors (2016-present)
AIPL1 is a specialized chaperone of the visual effector enzyme phosphodiesterase-6 (PDE6). Mutations in AIPL1 cause Leber congenital amaurosis type 4, a severe form of childhood blindness. We have examined the effects of mutations in the C-terminal region of human AIPL1 (hAIPL1) on the protein structure and function in order to ascertain the mechanism and/or the likelihood of disease causation.
hAIPL1, C-terminally truncated hAIPL1(1-316), R302L and P376S mutants were purified following expression in E. coli. The secondary structure and thermal stability of the proteins were examined with circular dichroism. Solution structures were probed by static light scattering and Small Angle X-ray Scattering (SAXS), and low resolution models were derived based on the SAXS data. Binding of farnesylated C-terminal peptide of PDE6A to hAIPL1 proteins was assessed using FRET.
Deletion of the C-terminal region or the R302L and P376S mutations did not appreciably alter the secondary structure or the thermal stability of hAIPL1. hAIPL1, hAIPL1(1-316), R302L and P376S behaved as monomeric proteins with little or no propensity for dimerization. SAXS analyses indicated that the C-terminal region of hAIPL1 assumes a relatively extended conformation and does not appear to interact with the rest of the molecule. Compared to hAIPL1, hAIPL1(1-316), R302L and P376S displayed very modest reductions in affinity to farnesylated PDE6A peptide.
The C-terminal region of hAIPL1 apparently has no influence on the FKBP and TPR-domains of the protein. Our results are consistent with the notion that R302L and P376S are benign variants rather than disease causing mutants of hAIPL1.
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