Structural basis of the P4B ATPase lipid flippase activity

Lin Bai,H Diessel Duan,Qinglong You,Bhawik K Jain,Todd R Graham,Mehmet Takar,Huilin Li

doi:10.1038/s41467-021-26273-0

Lin Bai, H Diessel Duan + Show 5 more

Open Access

https://doi.org/10.1038/s41467-021-26273-0

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Abstract

P4 ATPases are lipid flippases that are phylogenetically grouped into P4A, P4B and P4C clades. The P4A ATPases are heterodimers composed of a catalytic α-subunit and accessory β-subunit, and the structures of several heterodimeric flippases have been reported. The S. cerevisiae Neo1 and its orthologs represent the P4B ATPases, which function as monomeric flippases without a β-subunit. It has been unclear whether monomeric flippases retain the architecture and transport mechanism of the dimeric flippases. Here we report the structure of a P4B ATPase, Neo1, in its E1-ATP, E2P-transition, and E2P states. The structure reveals a conserved architecture as well as highly similar functional intermediate states relative to dimeric flippases. Consistently, structure-guided mutagenesis of residues in the proposed substrate translocation path disrupted Neo1’s ability to establish membrane asymmetry. These observations indicate that evolutionarily distant P4 ATPases use a structurally conserved mechanism for substrate transport.

Highlights

P4 ATPases are lipid flippases that are phylogenetically grouped into P4A, P4B and P4C clades
Most P-type ATPases have a conserved architecture consisting of a transmembrane domain (TMD), an actuator domain (A-domain), a nucleotide-binding domain (N-domain), and a phosphorylation domain (P-domain)[12]
Neo[1], Mon[2], Arl[1], and Dop[1] assemble into a membrane remodeling complex[40,41]. These regulatory proteins are crucial for yeast growth and likely interact with Neo[1] only transiently with low binding affinity, but little is known about their functions

Summary

Introduction

P4 ATPases are lipid flippases that are phylogenetically grouped into P4A, P4B and P4C clades. It has been unclear whether the monomeric lipid flippase Neo[1] functions by a similar mechanism or whether the cytosolically exposed exit gate is a conserved aspect of the substrate translocation path for P4 ATPases that lack a β-subunit. The conformation and the substrate transport path of Neo[1] in the E2P state highly resemble those of the Drs[2], Dnf[1], ATP8A1, and ATP11C flippases in the same E2P state (Supplementary Fig. 6).

Results

Conclusion