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Cyclization of the intrinsically disordered a1Sdihydropyridine receptor II-III loop enhances secondary structure and in vitro function

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posted on 2024-11-15, 02:15 authored by Han Shen TaeHan Shen Tae, Yanfang Cui, Yamuna Karunasekara, Philip G Board, Angela Dulhunty, Marco G Casarotto
A key component of excitation contraction (EC) coupling in skeletal muscle is the cytoplasmic linker (II-III loop) between the second and third transmembrane repeats of the α1S subunit of the dihydropyridine receptor (DHPR). The II-III loop has been previously examined in vitro using a linear II-III loop with unrestrained N- and C-terminal ends. To better reproduce the loop structure in its native environment (tethered to the DHPR transmembrane domains), we have joined the N and C termini using intein-mediated technology. Circular dichroism and NMR spectroscopy revealed a structural shift in the cyclized loop toward a protein with increased α-helical and β-strand structure in a region of the loop implicated in its in vitro function and also in a critical region for EC coupling. The affinity of binding of the II-III loop binding to the SPRY2 domain of the skeletal ryanodine receptor (RyR1) increased 4-fold, and its ability to activate RyR1 channels in lipid bilayers was enhanced 3-fold by cyclization. These functional changes were predicted consequences of the structural enhancement. We suggest that tethering the N and C termini stabilized secondary structural elements in the DHPR II-III loop and may reflect structural and dynamic characteristics of the loop that are inherent in EC coupling.

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Citation

Tae, H., Cui, Y., Karunasekara, Y., Board, P. G., Dulhunty, A. F. & Casarotto, M. G. (2011). Cyclization of the intrinsically disordered a1Sdihydropyridine receptor II-III loop enhances secondary structure and in vitro function. Journal of Biological Chemistry, 286 (25), 22589-22599.

Journal title

Journal of Biological Chemistry

Volume

286

Issue

25

Pagination

22589-22599

Language

English

RIS ID

120849

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