Crystal structure of UbiX, an aromatic acid decarboxylase from the psychrophilic bacterium Colwellia psychrerythraea that undergoes FMN-induced conformational changes.

Hackwon Do,Jun Hyuck Lee,Soo Jin Kim,Chang Woo Lee,Hyun Park,Hyun Ho Park,Hajeung Park,Han-Woo Kim,Ho Min Kim

doi:10.1038/srep08196

Abstract

The ubiX gene of Colwellia psychrerythraea strain 34H encodes a 3-octaprenyl-4-hydroxybenzoate carboxylase (CpsUbiX, UniProtKB code: Q489U8) that is involved in the third step of the ubiquinone biosynthesis pathway and harbors a flavin mononucleotide (FMN) as a potential cofactor. Here, we report the crystal structures of two forms of CpsUbiX: an FMN-bound wild type form and an FMN-unbound V47S mutant form. CpsUbiX is a dodecameric enzyme, and each monomer possesses a typical Rossmann-fold structure. The FMN-binding domain of UbiX is composed of three neighboring subunits. The highly conserved Gly15, Ser41, Val47, and Tyr171 residues play important roles in FMN binding. Structural comparison of the FMN-bound wild type form with the FMN-free form reveals a significant conformational difference in the C-terminal loop region (comprising residues 170–176 and 195–206). Subsequent computational modeling and liposome binding assay both suggest that the conformational flexibility observed in the C-terminal loops plays an important role in substrate and lipid bindings. The crystal structures presented in this work provide structural framework and insights into the catalytic mechanism of CpsUbiX.

Highlights

Crystal structure of UbiX, an aromatic acid decarboxylase from the psychrophilic bacterium Colwellia psychrerythraea that undergoes
The ubiX gene of Colwellia psychrerythraea strain 34H encodes a 3-octaprenyl-4-hydroxybenzoate carboxylase (CpsUbiX, UniProtKB code: Q489U8) that is involved in the third step of the ubiquinone biosynthesis pathway and harbors a flavin mononucleotide (FMN) as a potential cofactor
FMN-bound Colwellia psychrerythraea 34H UbiX protein (CpsUbiX) crystallized in the orthorhombic space group (C2221) with 6 monomers in the asymmetric unit, while FMN-free CpsUbiX crystallized in the cubic space group (P23) with one molecule in the asymmetric unit

Summary

Introduction

Crystal structure of UbiX, an aromatic acid decarboxylase from the psychrophilic bacterium Colwellia psychrerythraea that undergoes. Previous studies have shown that the expression of these two proteins is regulated by growth conditions and that both enzymes are required for the normal production of ubiquinone in the log phase of Escherichia coli cells[4] Despite their biologically important functions in the electron transport system, limited information is available regarding their substrate recognition, specificity, and reaction mechanisms. This is partly because the native substrates of these proteins, the membrane-associated hydrophobic polyprenyl p-hydroxybenzoate (PPPHB), are water-insoluble, and this impedes in vitro enzyme activity assays as well as determination of the crystal structures of the proteins in complex with the substrates. We propose that the C-terminal region plays the role of a gatekeeper for FMN and substrate binding

Methods

Results

Conclusion