Modulation of human 5-lipoxygenase activity by membrane lipids

    Authors

    A. H. Pande; D. Moe; K. N. Nemec; S. Qin; S. H. Tan;S. A. Tatulian

    Comments

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    Abbreviated Journal Title

    Biochemistry

    Keywords

    TRANSFORM INFRARED-SPECTROSCOPY; CYTOSOLIC PHOSPHOLIPASE A(2); HUMAN; ALVEOLAR MACROPHAGES; BASOPHILIC LEUKEMIA-CELLS; BETA-BARREL DOMAIN; N-TERMINAL DOMAIN; ARACHIDONATE 5-LIPOXYGENASE; SOYBEAN LIPOXYGENASE; 3-DIMENSIONAL STRUCTURE; LEUKOTRIENE SYNTHESIS; Biochemistry & Molecular Biology

    Abstract

    Mammalian 5-lipoxygenase (5-LO) catalyzes the conversion of arachidonic acid (AA) to leukotrienes, potent inflammatory mediators. 5-LO is activated by a Ca2+-mediated translocation to membranes, and dernonstrates the characteristic features of interfacially activated enzymes, yet the mechanism of membrane binding of 5-LO is not well understood. In an attempt to understand the mechanism of lipid-mediated activation of 5-LO, we have studied the effects of a large set of lipids on human recombinant 5-LO activity, as well as mutual structural effects of 5-LO and membranes. In the presence of 0.35 mM phosphatidyleholine (PC) and 0.2 mM Ca2+, there was substrate inhibition at > 100 muM AA. Data analysis at low AA concentrations yielded the following: K-m approximate to 103 muM and k(cat) approximate to 56 s(-1). 5-LO activity was Supported by PC more than by any other lipid tested except for a cationic lipid, which was more stimulatory than PC. Binding of 5-LO to zwitterionic and acidic membranes was relatively weak; the extent of binding increased 4-8 times in the presence of Ca2+, whereas binding to cationic membranes was stronger and essentially Ca2+-independent. Polarized attenuated total reflection infrared experiments implied that 5-LO binds to membranes at a defined orientation with the symmetry axis of the putative N-terminal beta-barrel tilted similar to45degrees from the membrane normal. Furthermore, membrane binding of 5-LO resulted in dehydration of the membrane surface and was paralleled with stabilization of the structures of both 5-LO and the membrane. Our results provide insight into the understanding of the effects of membrane surface properties on 5-LO-membrane interactions and the interfacial activation of 5-LO.

    Journal Title

    Biochemistry

    Volume

    43

    Issue/Number

    46

    Publication Date

    1-1-2004

    Document Type

    Article

    Language

    English

    First Page

    14653

    Last Page

    14666

    WOS Identifier

    WOS:000225172800016

    ISSN

    0006-2960

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