Purification, characterization and crystallization of the F-ATPase from Paracoccus denitrificans.

Abstract:

:The structures of F-ATPases have been determined predominantly with mitochondrial enzymes, but hitherto no F-ATPase has been crystallized intact. A high-resolution model of the bovine enzyme built up from separate sub-structures determined by X-ray crystallography contains about 85% of the entire complex, but it lacks a crucial region that provides a transmembrane proton pathway involved in the generation of the rotary mechanism that drives the synthesis of ATP. Here the isolation, characterization and crystallization of an integral F-ATPase complex from the α-proteobacterium Paracoccus denitrificans are described. Unlike many eubacterial F-ATPases, which can both synthesize and hydrolyse ATP, the P. denitrificans enzyme can only carry out the synthetic reaction. The mechanism of inhibition of its ATP hydrolytic activity involves a ζ inhibitor protein, which binds to the catalytic F₁-domain of the enzyme. The complex that has been crystallized, and the crystals themselves, contain the nine core proteins of the complete F-ATPase complex plus the ζ inhibitor protein. The formation of crystals depends upon the presence of bound bacterial cardiolipin and phospholipid molecules; when they were removed, the complex failed to crystallize. The experiments open the way to an atomic structure of an F-ATPase complex.

journal_name

Open Biol

journal_title

Open biology

authors

Morales-Rios E,Watt IN,Zhang Q,Ding S,Fearnley IM,Montgomery MG,Wakelam MJ,Walker JE

doi

10.1098/rsob.150119

subject

Has Abstract

pub_date

2015-09-01 00:00:00

pages

150119

issue

9

issn

2046-2441

pii

rsob.150119

journal_volume

5

pub_type

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