A conserved pi-cation and an electrostatic bridge are essential for 11R-lipoxygenase catalysis and structural stability

作者：Eek Priit; Piht Mari Ann; Raetsep Margus; Freiberg Arvi; Jaerving Ivar; Samel Nigulas

来源：Biochimica et Biophysica Acta-Molecular and Cell Biology of Lipids, 2015, 1851(10): 1377-1382.

DOI：10.1016/j.bbalip.2015.07.007

• Lipoxygenase
• Allosteric regulation
• Domain interaction
• PLAT domain
• Arachidonic acid
• Thermal denaturation

Lipoxygenases (LOXs) are lipid-peroxidizing enzymes that consist of a regulatory calcium- and membranebinding PLAT (polycystin-1, lipoxygenase, alpha-toxin) domain and a catalytic domain. In a previous study, the crystal structure of an 11R-LOX revealed a conserved pi-cation bridge connecting these two domains which could mediate the regulatory effect of the PLAT domain to the active site. Here we analyzed the role of residues Trpl 07 and Lys172 that constitute the pi-cation bridge in 11R-LOX along with Arg106 and Asp173-a potential salt bridge, which could also contribute to the inter-domain communication. According to our kinetic assays and protein unfolding experiments conducted using differential scanning fluorimetry and circular dichroism spectroscopy, mutants with a disrupted link display diminished catalytic activity alongside reduced stability of the protein fold. The results demonstrate that both these bridges contribute to the two-domain interface, and are important for proper enzyme activation.

• 出版日期2015-10