The Korean Journal of Physiology and Pharmacology

The Korean Society of Pharmacology (대한약리학회)
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Identification of Lys49-PLA2 from crude venom of Crotalus atrox as a human neutrophil-calcium modulating protein

  • Sultan, Md. Tipu (Department of Pharmacology, Institute of Natural Medicine, College of Medicine, Hallym University) ;
  • Li, Hong-Mei (Department of Pharmacology, Institute of Natural Medicine, College of Medicine, Hallym University) ;
  • Lee, Yong Zu (ProteomeTech Inc.) ;
  • Lim, Soon Sung (Department of Food Science and Nutrition, Hallym University) ;
  • Song, Dong-Keun (Department of Pharmacology, Institute of Natural Medicine, College of Medicine, Hallym University)
  • Received : 2015.09.17
  • Accepted : 2016.01.03
  • Published : 2016.03.01
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Abstract

We fortuitously observed a human neutrophil intracellular free-calcium concentration ($[Ca^{2+}]_i$) increasing activity in the commercially available phosphodiesterase I (PDE I), which is actually dried crude venom of Crotalus atrox. As this activity was not observed with another commercially available pure PDE I, we tried to find out the causative molecule(s) present in 'crude' PDE, and identified Lys49-phospholipase A2 (Lys49-PLA2 or K49-PLA2), a catalytically inactive protein which belongs to the phospholipase A2 family, by activity-driven three HPLC (reverse phase, size exclusion, reverse phase) steps followed by SDS-PAGE and LC-MS/MS. K49-PLA2 induced $Ca^{2+}$ influx in human neutrophils without any cytotoxic effect. Two calcium channel inhibitors, 2-aminoetoxydiphenyl borate (2-APB) ($30{\mu}M$) and SKF-96365 ($20{\mu}M$) significantly inhibited K49-PLA2-induced $[Ca^{2+}]_i$ increase. These results suggest that K49-PLA2 modulates $[Ca^{2+}]_i$ in human neutrophils via 2-APB- and SKF-96365-sensitive calcium channels without causing membrane disruption.

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