Calcium modulates membrane association, positional specificity, and product distribution in dual positional specific maize lipoxygenase-1

• Asp38, Glu127 and Glu201 were identified as putative calcium binding residues in maize lipoxygenase-1.
• Asp38 and Leu37 were not essential for membrane association of maize lipoxygenase-1.
• Calcium promotes the oxidation of linoleic acid by binding to maize lipoxygenase-1.
• Calcium interacts with substrate and regulates the access to the maize lipoxygenase-1.
• Calcium modulates maize lipoxygenase-1 reaction favoring 13-isomeric and kinetically controlled products.

This study investigates how calcium modulates the properties of dual positional specific maize lipoxygenase-1, including its interaction with substrate, association with subcellular membrane and alteration of product distribution. Bioinformatic analyses identified Asp38, Glu127 and Glu201 as putative calcium binding residues and Leu37 as a flanking hydrophobic residue also potentially involved in calcium-mediated binding of the enzyme to subcellular membranes. Asp38 and Leu37 were shown to be important but not essential for calcium-mediated association of maize lipoxygenase-1 to subcellular membranes in vitro. Kinetic studies demonstrate that catalytic efficiency (Vmax/Km) shows a bell-shaped dependence on log of the molar ratio of substrate to unbound calcium. Calcium also modulates product distribution of the maize lipoxygenase-1 reaction, favoring 13-positional specificity and increasing the relative amount of (E,Z)-isomeric products. The results suggest that calcium regulates the maize lipoxygenase-1 reaction by binding to substrate, and by promoting binding of substrate to enzyme and association of maize lipoxygenase-1 to subcellular membranes.

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