1huv

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spinqualitylabelsall models 1huv, resolution 2.15Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

CRYSTAL STRUCTURE OF A SOLUBLE MUTANT OF THE MEMBRANE-ASSOCIATED (S)-MANDELATE DEHYDROGENASE FROM PSEUDOMONAS PUTIDA AT 2.15A RESOLUTION

Overview

The structure of an active mutant of (S)-mandelate dehydrogenase, (MDH-GOX2) from Pseudomonas putida has been determined at 2.15 A, resolution. The membrane-associated flavoenzyme (S)-mandelate, dehydrogenase (MDH) catalyzes the oxidation of (S)-mandelate to give a, flavin hydroquinone intermediate which is subsequently reoxidized by an, organic oxidant residing in the membrane. The enzyme was rendered soluble, by replacing its 39-residue membrane-binding peptide segment with a, corresponding 20-residue segment from its soluble homologue, glycolate, oxidase (GOX). Because of their amphipathic nature and peculiar, solubilization properties, membrane proteins are notoriously difficult to, crystallize, yet represent a large fraction of the proteins encoded by, genomes currently being deciphered. Here we present the first report of, such a structure in which an internal membrane-binding segment has been, replaced, leading to successful crystallization of the fully active enzyme, in the absence of detergents. This approach may have general application, to other membrane-bound proteins. The overall fold of the molecule is that, of a TIM barrel, and it forms a tight tetramer within the crystal lattice, that has circular 4-fold symmetry. The structure of MDH-GOX2 reveals how, this molecule can interact with a membrane, although it is limited by the, absence of a membrane-binding segment. MDH-GOX2 and GOX adopt similar, conformations, yet they retain features characteristic of membrane and, globular proteins, respectively. MDH-GOX2 has a distinctly electropositive, surface capable of interacting with the membrane, while the opposite, surface is largely electronegative. GOX shows no such pattern. MDH appears, to form a new class of monotopic integral membrane protein that interacts, with the membrane through coplanar electrostatic binding surfaces and, hydrophobic interactions, thus combining features of both the, prostaglandin synthase/squaline-hopine cyclase and the C-2 coagulation, factor domain classes of membrane proteins.

About this Structure

1HUV is a Single protein structure of sequence from Pseudomonas putida with SO4, FMN and MES as ligands. Full crystallographic information is available from OCA.

Reference

Structure of an active soluble mutant of the membrane-associated (S)-mandelate dehydrogenase., Sukumar N, Xu Y, Gatti DL, Mitra B, Mathews FS, Biochemistry. 2001 Aug 21;40(33):9870-8. PMID:11502180

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