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Inhibition of secreted phospholipase A2 by proanthocyanidins: a comparative enzymological and in silico modeling study.

ABSTRACT: Secreted phospholipase A2 (PLA2) plays a critical role in mobilizing arachidonic acid in phospholipids. We have previously reported that PLA2 is inhibited by B-type proanthocyanidins (PaCs). To further understand the inhibitory activity of these compounds, we compared the inhibitory potency of B-type PaCs to that of A-type PaCs and modeled them with PLA2 using in silico techniques. The B-type trimer and tetramer inhibited PLA2 (IC50 = 16 and 10 ?M). The A-type compounds were less potent (18-35% inhibition at 50 ?M). The active site of PLA2 lies in a hydrophobic tunnel. Modeling studies revealed that the B-type PaCs occupy this tunnel and are stabilized by a number of van der Waals interactions. The result is reduced substrate access to the active site. The A-type compounds can occupy this tunnel only by shifting the N-terminal loop outward. Our data provide a structural basis to screen additional PaCs for anti-PLA2 activity.

SUBMITTER: Lambert JD

PROVIDER: S-EPMC3740037 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature

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Inhibition of secreted phospholipase A2 by proanthocyanidins: a comparative enzymological and in silico modeling study.

Lambert Joshua D JD Yennawar Neela N Gu Yeyi Y Elias Ryan J RJ

Journal of agricultural and food chemistry 20120718 30

Secreted phospholipase A2 (PLA2) plays a critical role in mobilizing arachidonic acid in phospholipids. We have previously reported that PLA2 is inhibited by B-type proanthocyanidins (PaCs). To further understand the inhibitory activity of these compounds, we compared the inhibitory potency of B-type PaCs to that of A-type PaCs and modeled them with PLA2 using in silico techniques. The B-type trimer and tetramer inhibited PLA2 (IC50 = 16 and 10 μM). The A-type compounds were less potent (18-35% ...[more]

PMID: 22779786

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Project description:BackgroundPhospholipase A2s mediate the rate-limiting step in the formation of eicosanoids such as cysteinyl leukotrienes (CysLTs). Group IVA cytosolic PLA2α (cPLA2α) is thought to be the dominant PLA2 in eosinophils; however, eosinophils also have secreted PLA2 (sPLA2) activity that has not been fully defined.ObjectivesTo examine the expression of sPLA2 group X (sPLA2-X) in eosinophils, the participation of sPLA2-X in the formation of CysLTs, and the mechanism by which sPLA2-X initiates the synthesis of CysLTs in eosinophils.MethodsPeripheral blood eosinophils were obtained from volunteers with asthma and/or allergy. A rabbit polyclonal anti-sPLA2-X antibody identified sPLA2-X by Western blot. We used confocal microscopy to colocalize the sPLA2-X to intracellular structures. An inhibitor of sPLA2-X (ROC-0929) that does not inhibit other mammalian sPLA2s, as well as inhibitors of the mitogen-activated kinase cascade (MAPK) and cPLA2α, was used to examine the mechanism of N-formyl-methionyl-leucyl-phenylalanine (fMLP)-mediated formation of CysLT.ResultsEosinophils express the mammalian sPLA2-X gene (PLA2G10). The sPLA2-X protein is located in the endoplasmic reticulum, golgi, and granules of eosinophils and moves to the granules and lipid bodies during fMLP-mediated activation. Selective sPLA2-X inhibition attenuated the fMLP-mediated release of arachidonic acid and CysLT formation by eosinophils. Inhibitors of p38, extracellular-signal-regulated kinases 1/2 (p44/42 MAPK), c-Jun N-terminal kinase, and cPLA2α also attenuated the fMLP-mediated formation of CysLT. The sPLA2-X inhibitor reduced the phosphorylation of p38 and extracellular-signal-regulated kinases 1/2 (p44/42 MAPK) as well as cPLA2α during cellular activation, indicating that sPLA2-X is involved in activating the MAPK cascade leading to the formation of CysLT via cPLA2α. We further demonstrate that sPLA2-X is activated before secretion from the cell during activation. Short-term priming with IL-13 and TNF/IL-1β increased the expression of PLA2G10 by eosinophils.ConclusionsThese results demonstrate that sPLA2-X plays a significant role in the formation of CysLTs by human eosinophils. The predominant role of the enzyme is the regulation of MAPK activation that leads to the phosphorylation of cPLA2α. The sPLA2-X protein is regulated by proteolytic cleavage, suggesting that an inflammatory environment may promote the formation of CysLTs through this mechanism. These results have important implications for the treatment of eosinophilic disorders such as asthma.

Dataset Information

Inhibition of secreted phospholipase A2 by proanthocyanidins: a comparative enzymological and in silico modeling study.

Publications

Inhibition of secreted phospholipase A2 by proanthocyanidins: a comparative enzymological and in silico modeling study.

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