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

Lopez-Redondo, Maria; Fan, Shujie; Koide, Akiko; Koide, Shohei; Beckstein, Oliver; Stokes, David L.

doi:10.1085/jgp.202112873

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

Journal Article · Tue Jul 13 00:00:00 EDT 2021 · Journal of General Physiology

DOI:https://doi.org/10.1085/jgp.202112873· OSTI ID:1903935

Lopez-Redondo, Maria ^[1];

^[2];

^[1];

^[2];

^[1]

New York University School of Medicine, New York, NY (United States)
Arizona State University, Tempe, AZ (United States)

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.

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Research Organization:: New York University School of Medicine, New York, NY (United States); Arizona State Univ., Tempe, AZ (United States)

Sponsoring Organization:: 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)

Grant/Contract Number:: AC05-76RL01830; 1R01GM125081; 754513; U24GM129547; ACI-1548562

OSTI ID:: 1903935

Journal Information:: Journal of General Physiology, Vol. 153, Issue 8; ISSN 0022-1295

Publisher:: Rockefeller University PressCopyright Statement

Country of Publication:: United States

Language:: English

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

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