A Covalent Linker Allows for Membrane Targeting of an Oxylipin Biosynthetic Complex

A naturally occurring bifunctional protein from Plexaura homomalla links sequential catalytic activities in an oxylipin biosynthetic pathway. The C-terminal lipoxygenase (LOX) portion of the molecule catalyzes the transformation of arachidonic acid (AA) to the corresponding 8R-hydroperoxide, and the...

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Veröffentlicht in:Biochemistry (Easton) 2008-10, Vol.47 (40), p.10665-10676
Hauptverfasser: Gilbert, Nathaniel C, Niebuhr, Marc, Tsuruta, Hiro, Bordelon, Tee, Ridderbusch, Oswin, Dassey, Adam, Brash, Alan R, Bartlett, Sue G, Newcomer, Marcia E
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Sprache:eng
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Zusammenfassung:A naturally occurring bifunctional protein from Plexaura homomalla links sequential catalytic activities in an oxylipin biosynthetic pathway. The C-terminal lipoxygenase (LOX) portion of the molecule catalyzes the transformation of arachidonic acid (AA) to the corresponding 8R-hydroperoxide, and the N-terminal allene oxide synthase (AOS) domain promotes the conversion of the hydroperoxide intermediate to the product allene oxide (AO). Small-angle X-ray scattering data indicate that in the absence of a covalent linkage the two catalytic domains that transform AA to AO associate to form a complex that recapitulates the structure of the bifunctional protein. The SAXS data also support a model for LOX and AOS domain orientation in the fusion protein inferred from a low-resolution crystal structure. However, results of membrane binding experiments indicate that covalent linkage of the domains is required for Ca2+-dependent membrane targeting of the sequential activities, despite the noncovalent domain association. Furthermore, membrane targeting is accompanied by a conformational change as monitored by specific proteolysis of the linker that joins the AOS and LOX domains. Our data are consistent with a model in which Ca2+-dependent membrane binding relieves the noncovalent interactions between the AOS and LOX domains and suggests that the C2-like domain of LOX mediates both protein−protein and protein−membrane interactions.
ISSN:0006-2960
1520-4995
DOI:10.1021/bi800751p