An Arginine-Rich Loop is Critical for the Modulation of the Water Permeability of Aquaporin 0

Abstract

Aquaporin 0 (AQP0), the major intrinsic protein of the eye lens, plays a vital role in maintaining lens clarity by facilitating water transport. AQP0 reduces its osmotic water permeability (Pf) in response to increases in the external calcium concentration, an effect that is mediated through an interaction with calmodulin (CaM). Calcium sensitivity of AQP0 Pf is further modified by serine phosphorylation. Despite recent structural characterization of the AQP0-CaM complex, the mechanism used by CaM to modulate AQP0 remains poorly understood. We employed a combination of Brownian and molecular dynamics simulations to identify the critical features of the AQP0-CaM interaction. Brownian dynamics (BD) simulations suggest that serine phosphorylation of AQP0 does not significantly reduce CaM-binding to the whole AQP0 protein, in contrast to the experimental observation that phosphorylation does significantly reduce binding to C-terminus AQP0 peptides. Comparative MD simulation studies show that AQP0 phosphorylation changes contacts between AQP0 and CaM, particularly at a small arginine-rich loop on the AQP0 cytosolic face. This charged loop allosterically couples CaM to the second constriction site residues of AQP0 through an interaction with R156. Additionally, we observe that R153 increases the size of the pore opening through an interaction with the Y149 hydroxyl group, which is necessary for maintaining high permeability states of AQP0. Experimental and simulation data support the notion that serine phosphorylation of AQP0 changes the calcium sensitivity of Pf by modifying CaM contacts with the cytosolic arginine-rich loop rather than by inhibiting CaM-binding.