Schenk, Gerhard and Neidig, Michael L. and Zhou, Jing and Holman, Theodore R. and Solomon, Edward I.
Spectroscopic Characterization of Soybean Lipoxygenase-1 Mutants: the Role of
Second Coordination Sphere Residues in the Regulation of Enzyme Activity.
Biochemistry, 42 (24).
Lipoxygenases are non-heme iron enzymes, which catalyze the stereo- and regiospecific
hydroperoxidation of unsaturated fatty acids. Spectroscopic studies on soybean lipoxygenase have shown
that the ferrous form of the enzyme is a mixture of five- and six-coordinate species (40 and 60%,
respectively). Addition of substrate leads to a purely six-coordinate form. A series of mutations in the
second coordination sphere (Q697E, Q697N, Q495A, and Q495E) were generated, and the structures of
the mutants were solved by crystallography [Tomchick et al. (2001) Biochemistry 40, 7509-7517]. While
this study clearly showed the contribution of H-bond interactions between the first and the second
coordination spheres in catalysis, no correlation with the coordination environment of the FeII was observed.
A recent study using density-functional theory [Lehnert and Solomon (2002) J. Biol. Inorg. Chem. 8,
294-305] indicated that coordination flexibility, involving the Asn694 ligand, is regulated via H-bond
interactions. In this paper, we investigate the solution structures of the second coordination sphere mutants
using CD and MCD spectroscopy since these techniques are more sensitive indicators of the first
coordination sphere ligation of FeII systems. Our data demonstrate that the iron coordination environment
directly relates to activity, with the mutations that have the ability to form a five-coordinate/six-coordinate
mixture being more active. We propose that the H-bond between the weak Asn694 ligand and the Gln697
plays a key role in the modulation of the coordination flexibility of Asn694, and thus, is crucial for the
regulation of enzyme reactivity
||Spectroscopic Characterization; Soybean Lipoxygenase-1 Mutants; Sphere Residues; Enzyme Activity;
||Faculty of Science and Engineering > Chemistry
||30 May 2012 09:28
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||American Chemical Society
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