Structure of a monotropic membrane enzyme
Michael Malkowski group (Hauptman-Woodward Medical Research Institute and SUNY - Buffalo) and collaborators
The group of Michael Malkowski at the Hauptman-Woodward
Medical Research Institute and Department of Structural Biology, SUNY at
Buffalo, determined the x-ray crystal structure of Arabidopsis thaliana
alpha-dioxygenase (alpha-DOX) to 1.5-Å resolution using MAD phasing,
which yielded insight into diversity with the cyclooxygenase-peroxidase
superfamily of monotopic membrane enzymes. alpha-DOX is a heme-containing
enzyme that catalyzes the oxygenation of fatty acids into 2(R)-hydroperoxide
products via the stereospecific removal of the pro-R hydrogen from carbon-2
of the fatty acid substrate. Despite the low level of sequence identity,
alpha-DOX shares common catalytic features with cyclooxygenase (COX) within
the cyclooxygenase-peroxidase superfamily, including the use of a tyrosyl
radical during catalysis. The overall structure of alpha-DOX is
predominantly alpha-helical in nature and comprised of a base domain and a
catalytic domain. The base domain exhibits a low degree of structural
homology with the membrane-binding domain of COX. The catalytic domain shows
the highest degree of similarity with COX, where 21 of the 22 alpha-helical
elements are conserved. Four alpha-helices form the heme-binding cleft and
walls of the active site channel, which are highly similar to the
alpha-helices that define the cyclooxygenase channel in COX. These four
alpha-helices contain the proximal and distal histidines required for heme
binding, the catalytic tyrosine, as well as highly conserved arginine and
histidine residues involved in fatty acid binding and positioning. Unlike
COX, the heme-binding cleft is buried deep within the catalytic domain of the
enzyme, where access to the distal face of the heme is restricted by the
presence of two extended inserts unique to alpha-DOX. The alpha-DOX
structure represents the first member of the alpha-DOX subfamily to be
structurally characterized within the cyclooxygenase-peroxidase family of
heme-containing proteins.
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Figure: Comparison of the structures
of COX and alpha-DOX. (Left) Ribbon diagram of mouse COX-2 (PDB id 3HS5) with
arachidonic acid (yellow) bound within the cyclooxygenase channel. The
alpha-helices of the membrane-binding domain are colored in gold. (Right)
Ribbon diagram of alpha-DOX. The alpha-helices of the base domain are colored
in blue. The heme moiety in each structure is colored red. |
Citation:
Goulah, CC, Zhu, G, Koszelak-Rosenblum, M, Malkowski, MG. The Crystal
Structure of [alpha]-Dioxygenase Provides Insight into Diversity in the
Cyclooxygenase-Peroxidase Superfamily, Biochemistry 52, 1364-1372 (2013).
DOI: 10.1021/bi400013k
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