Title

Uncoupling of the phosphatase produces a deeper relaxation of Ci-VSP

ORCiD

Carlos A. Villalba-Galea: 0000-0002-6489-4651

Document Type

Abstract

Conference Title/Conference Publication

Biophysical Journal

Organization

Biophysical Society 53rd Annual Meeting

Location

Boston, MA

Conference Dates

February 28-March 4, 2009

Date of Presentation

2-28-2009

ISSN

0006-3495

Volume

96

Issue

3, Supplement 1

DOI

10.1016/j.bpj.2008.12.1992

First Page

370a

Abstract

The coupling of the Voltage sensing domain (VSD) and the phosphatase domain (PD) of the Ciona intestinalis voltage sensing phosphatase (Ci-VSP) is mediated by a putative Phospholipid Biding Motif (PBM), located between the two domains. During depolarization, the movement of the S4 favors the binding of the PBM to the membrane, placing the PD in position to carry out its function. We have shown that upon prolonged depolarization, the VSD of Ci-VSP evolves to a relaxed state, as shown by a shift of the Q-V curve (Villalba-Galea et al., 2008). As a consequence of the relaxation of the VSD, fluorescence changes of a probe attached to the extracellular end of the Ci-VSP S4 were observed. However, we have now found that the relaxation produces changes along the entire S4 segment. We report here that mutations in the PBM, R253A-R254A, uncouple the sensing currents from the phosphatase activity, suggesting that the mutations abolish the coupling by disrupting the binding of the Arginine of the PBM to the phospholipid of the membrane. Interestingly, these mutations also increase the rate of the sensing currents during repolarization. A simple interpretation is that the S4 movement is restricted by the binding of the PBM to the membrane. If this is the case, it may also restrict the relaxation such that during a long depolarization the S4 could undergo a larger movement and reach a deeper relaxed state. Consistent with this interpretation we found that in the PBM mutant, the shift of the Q-V curve during prolonged depolarization is enhanced while the recovery time from the relaxed state is increased. Supported by NIHGM30376

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