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Title: Zinc binding alters the conformational dynamics and drives the transport cycle of the cation diffusion facilitator YiiP

Abstract

YiiP is a secondary transporter that couples Zn2+ transport to the proton motive force. Structural studies of YiiP from prokaryotes and Znt8 from humans have revealed three different Zn2+ sites and a conserved homodimeric architecture. These structures define the inward-facing and outward-facing states that characterize the archetypal alternating access mechanism of transport. To study the effects of Zn2+ binding on the conformational transition, we use cryo-EM together with molecular dynamics simulation to compare structures of YiiP from Shewanella oneidensis in the presence and absence of Zn2+. To enable single-particle cryo-EM, we used a phage-display library to develop a Fab antibody fragment with high affinity for YiiP, thus producing a YiiP/Fab complex. To perform MD simulations, we developed a nonbonded dummy model for Zn2+ and validated its performance with known Zn2+-binding proteins. Using these tools, we find that, in the presence of Zn2+, YiiP adopts an inward-facing conformation consistent with that previously seen in tubular crystals. After removal of Zn2+ with high-affinity chelators, YiiP exhibits enhanced flexibility and adopts a novel conformation that appears to be intermediate between inward-facing and outward-facing states. This conformation involves closure of a hydrophobic gate that has been postulated to control access to the primary transportmore » site. Comparison of several independent cryo-EM maps suggests that the transition from the inward-facing state is controlled by occupancy of a secondary Zn2+ site at the cytoplasmic membrane interface. This work enhances our understanding of individual Zn2+ binding sites and their role in the conformational dynamics that govern the transport cycle.« less

Authors:
 [1]; ORCiD logo [2]; ORCiD logo [1]; ORCiD logo [1]; ORCiD logo [2]; ORCiD logo [1]
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  1. New York University School of Medicine, New York, NY (United States)
  2. Arizona State University, Tempe, AZ (United States)
Publication Date:
Research Org.:
New York University School of Medicine, New York, NY (United States); Arizona State Univ., Tempe, AZ (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER); National Institutes of Health (NIH); European Union’s 2020; Marie Sklodowska-Curie Grant; Aarhus University Research Foundation; National Science Foundation (NSF)
OSTI Identifier:
1903935
Grant/Contract Number:  
AC05-76RL01830; 1R01GM125081; 754513; U24GM129547; ACI-1548562
Resource Type:
Accepted Manuscript
Journal Name:
Journal of General Physiology
Additional Journal Information:
Journal Volume: 153; Journal Issue: 8; Journal ID: ISSN 0022-1295
Publisher:
Rockefeller University Press
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; biophysics; membrane transport; protein structure and dynamics

Citation Formats

Lopez-Redondo, Maria, Fan, Shujie, Koide, Akiko, Koide, Shohei, Beckstein, Oliver, and Stokes, David L. Zinc binding alters the conformational dynamics and drives the transport cycle of the cation diffusion facilitator YiiP. United States: N. p., 2021. Web. doi:10.1085/jgp.202112873.
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Lopez-Redondo, Maria, Fan, Shujie, Koide, Akiko, Koide, Shohei, Beckstein, Oliver, & Stokes, David L. Zinc binding alters the conformational dynamics and drives the transport cycle of the cation diffusion facilitator YiiP. United States. https://doi.org/10.1085/jgp.202112873
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Lopez-Redondo, Maria, Fan, Shujie, Koide, Akiko, Koide, Shohei, Beckstein, Oliver, and Stokes, David L. Tue . "Zinc binding alters the conformational dynamics and drives the transport cycle of the cation diffusion facilitator YiiP". United States. https://doi.org/10.1085/jgp.202112873. https://www.osti.gov/servlets/purl/1903935.
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@article{osti_1903935,
title = {Zinc binding alters the conformational dynamics and drives the transport cycle of the cation diffusion facilitator YiiP},
author = {Lopez-Redondo, Maria and Fan, Shujie and Koide, Akiko and Koide, Shohei and Beckstein, Oliver and Stokes, David L.},
abstractNote = {YiiP is a secondary transporter that couples Zn2+ transport to the proton motive force. Structural studies of YiiP from prokaryotes and Znt8 from humans have revealed three different Zn2+ sites and a conserved homodimeric architecture. These structures define the inward-facing and outward-facing states that characterize the archetypal alternating access mechanism of transport. To study the effects of Zn2+ binding on the conformational transition, we use cryo-EM together with molecular dynamics simulation to compare structures of YiiP from Shewanella oneidensis in the presence and absence of Zn2+. To enable single-particle cryo-EM, we used a phage-display library to develop a Fab antibody fragment with high affinity for YiiP, thus producing a YiiP/Fab complex. To perform MD simulations, we developed a nonbonded dummy model for Zn2+ and validated its performance with known Zn2+-binding proteins. Using these tools, we find that, in the presence of Zn2+, YiiP adopts an inward-facing conformation consistent with that previously seen in tubular crystals. After removal of Zn2+ with high-affinity chelators, YiiP exhibits enhanced flexibility and adopts a novel conformation that appears to be intermediate between inward-facing and outward-facing states. This conformation involves closure of a hydrophobic gate that has been postulated to control access to the primary transport site. Comparison of several independent cryo-EM maps suggests that the transition from the inward-facing state is controlled by occupancy of a secondary Zn2+ site at the cytoplasmic membrane interface. This work enhances our understanding of individual Zn2+ binding sites and their role in the conformational dynamics that govern the transport cycle.},
doi = {10.1085/jgp.202112873},
journal = {Journal of General Physiology},
number = 8,
volume = 153,
place = {United States},
year = {Tue Jul 13 00:00:00 EDT 2021},
month = {Tue Jul 13 00:00:00 EDT 2021}
}
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