title: Functional Elucidation of Nemopilema nomurai and Cyanea nozakii Nematocyst Venoms' Lytic Activity Using Mass Spectrometry and Zymography.
abstract: Medusozoans utilize explosively discharging penetrant nematocysts to inject venom into prey. These venoms are composed of highly complex proteins and peptides with extensive bioactivities, as observed in vitro. Diverse enzymatic toxins have been putatively identified in the venom of jellyfish,Nemopilema nomuraiandCyanea nozakii, through examination of their proteomes and transcriptomes. However, functional examination of putative enzymatic components identified in proteomic approaches to elucidate potential bioactivities is critically needed.
  
  
  
  In this study, enzymatic toxins were functionally identified using a combined approach consisting ofin gelzymography and liquid chromatography tandem mass spectrometry (LC-MS/MS). The potential roles of metalloproteinases and lipases in hemolytic activity were explored using specific inhibitors.
  
  
  
  Zymography indicated that nematocyst venom possessed protease-, lipase- and hyaluronidase-class activities. Further, proteomic approaches using LC-MS/MS indicated sequence homology of proteolytic bands observed in zymography to extant zinc metalloproteinase-disintegrins and astacin metalloproteinases. Moreover, pre-incubation of the metalloproteinase inhibitor batimastat withN.nomurainematocyst venom resulted in an approximate 62% reduction of hemolysis compared to venom exposed sheep erythrocytes, suggesting that metalloproteinases contribute to hemolytic activity. Additionally, species within the molecular mass range of 14-18 kDa exhibited both egg yolk and erythrocyte lytic activities in gel overlay assays.
  
  
  
  For the first time, our findings demonstrate the contribution of jellyfish venom metalloproteinase and suggest the involvement of lipase species to hemolytic activity. Investigations of this relationship will facilitate a better understanding of the constituents and toxicity of jellyfish venom.
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. buqun08@163.com.|University of the Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100039, China. buqun08@163.com.
authors.forename: Yang
authors.initials: Y
authors.lastname: Yue
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. yuhuahua@qdio.ac.cn.
authors.forename: Huahua
authors.initials: H
authors.lastname: Yu
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. rongfengli@qdio.ac.cn.
authors.forename: Rongfeng
authors.initials: R
authors.lastname: Li
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. xingronge@qdio.ac.cn.
authors.forename: Ronge
authors.initials: R
authors.lastname: Xing
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. sliu@qdio.ac.cn.
authors.forename: Song
authors.initials: S
authors.lastname: Liu
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. lkc@qdio.ac.cn.
authors.forename: Kecheng
authors.initials: K
authors.lastname: Li
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. xueqinwang@qdio.ac.cn.
authors.forename: Xueqin
authors.initials: X
authors.lastname: Wang
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. chenxl@qdio.ac.cn.
authors.forename: Xiaolin
authors.initials: X
authors.lastname: Chen
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. pcli@qdio.ac.cn.
authors.forename: Pengcheng
authors.initials: P
authors.lastname: Li
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid; D003064:Cnidarian Venoms / Q000037:antagonists & inhibitors / Q000201:enzymology* / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006461:Hemolysis / Q000187:drug effects; D006821:Hyaluronoglucosaminidase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity; D008049:Lipase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity; D045726:Metalloproteases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000494:pharmacology; D040901:Proteomics / Q000379:methods*; D037401:Scyphozoa / Q000201:enzymology*; D034561:Sheep, Domestic; D053719:Tandem Mass Spectrometry
chemical_list: D003064:Cnidarian Venoms; D011480:Protease Inhibitors; D008049:Lipase; D006821:Hyaluronoglucosaminidase; D045726:Metalloproteases
keywords: Cyanea nozakii; LC-MS/MS; Nemopilema nomurai; PLA2 inhibitors; enzymatic toxins; hemolysis; jellyfish metalloproteinase inhibitors; zymography