title: Okinalysin, a novel P-I metalloproteinase from Ovophis okinavensis: biological properties and effect on vascular endothelial cells.
abstract: A novel hemorrhagic metalloproteinase, okinalysin, was isolated from the venom of Ovophis okinavensis. It possessed caseinolytic and hemorrhagic activities, and also hydrolyzed fibrinogen and collagen. These activities were inhibited by ethylenediaminetetraacetic acid (EDTA) but not by p-amidinophenyl methanesulfonyl fluoride hydrochloride (APMSF). The molecular mass of okinalysin was 22,202 Da measured by MALDI/TOF mass spectrometry. The primary structure of okinalysin was partially determined by Edman sequencing, and the putative zinc-binding domain HEXXHXXGXXH was found to be present in its structure. From these data, okinalysin is defined as a metalloproteinase belonging to a P-I class. The partial amino acid sequence of okinalysin was homologous to the C-terminus of MP 10, a putative metalloproteinase induced from transcriptome of the venom gland cDNA sequencing of O. okinavensis. Okinalysin possessed cytotoxic activity on cultured endothelial cells, and the EC50 on human pulmonary artery endothelial cells was determined to be 0.6 μg/mL. The histopathological study also showed that okinalysin causes the leakage of red blood cells and neutrophil infiltration. These results indicate that destruction of blood vessels by okinalysin is one of the main causes of hemorrhage.
authors.affiliation: Department of Microbiology, Faculty of Pharmacy, Meijo University, 150 Yagotoyama, Tenpaku, Nagoya 468-8503, Japan. ykomori@meijo-u.ac.jp.
authors.forename: Yumiko
authors.initials: Y
authors.lastname: Komori
authors.affiliation: Department of Microbiology, Faculty of Pharmacy, Meijo University, 150 Yagotoyama, Tenpaku, Nagoya 468-8503, Japan. paku.hgrn-ed15@ezweb.ne.jp.
authors.forename: Eri
authors.initials: E
authors.lastname: Murakami
authors.affiliation: Department of Microbiology, Faculty of Pharmacy, Meijo University, 150 Yagotoyama, Tenpaku, Nagoya 468-8503, Japan. kuchiya@meijo-u.ac.jp.
authors.forename: Kei-Ichi
authors.initials: K
authors.lastname: Uchiya
authors.affiliation: College of Pharmacy, Kinjo Gakuin University, 2-1723 Omori, Moriyama, Nagoya 463-8521, Japan. tunenono@kinjo-u.ac.jp.
authors.forename: Tunemasa
authors.initials: T
authors.lastname: Nonogaki
authors.affiliation: Department of Microbiology, Faculty of Pharmacy, Meijo University, 150 Yagotoyama, Tenpaku, Nagoya 468-8503, Japan. nikai@meijo-u.ac.jp.
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Nikai
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002364:Caseins / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D024141:Collagen Type IV / Q000378:metabolism; D003435:Crotalid Venoms / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism; D006470:Hemorrhage / Q000139:chemically induced; D006801:Humans; D006868:Hydrolysis; D045726:Metalloproteases / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000473:pathology; D011651:Pulmonary Artery; D013848:Thigh; D017819:Viperidae
chemical_list: D002364:Caseins; D024141:Collagen Type IV; D003435:Crotalid Venoms; D005340:Fibrinogen; D045726:Metalloproteases; C000597829:okinalysin protein, Ovophis okinavensis