Sea snake Hydrophis cyanocinctus venom. I. Purification, characterization and N-terminal sequence of two phospholipases A2.
Two phospholipases A2 (PLA2, H1 and H2) from sea snake Hydrophis cyanocinctus venom were purified to homogeneity in a single step using reversed-phase high performance liquid chromatography on a Nucleosil 7C18 column. The molecular weights of H1 and H2, as estimated by MALDI MS, were 13588.1 and 13247.2 Da, respectively. The N-terminal 60 amino acid residues were determined by direct automated Edman degradation analysis. Since both PLA2s show close sequence homologies to those of PLA2s from other Elapid snakes (60-84%) they have been tentatively classified as belonging to group-IA and Asp-49 phospholipases A2. Despite the sequence variation (18%) between H1 and H2, their general structural organization is very similar as shown by their clearly related CD spectra. Furthermore, both enzymes are quite thermostable (60-65 degrees C) as determined by temperature variable CD spectra, indicating that the enzymes contain compact folded structure, mainly based on the core structure of disulfide bridges. However, the major PLA2 (H1) shows higher toxicity to albino rats (LD50 i.p. 0.04 mg/kg) and purification resulted in 18-fold increase in toxicity over the crude or whole venom (LD50 i.p. 0.80 mg/kg). H1 also shows edema-inducing and indirect haemolytic but no haemorrhagic activity. Unlike the toxic PLA2-H1, enzyme H2 was not toxic to albino rats but showed edema-inducing and indirect haemolytic activities.
Ali
S A
SA
International Center for Chemical Sciences, HEJ Research Institute of Chemistry, University of Karachi, Pakistan. saali66@hotmail.com
Alam
J M
JM
Stoeva
S
S
Schütz
J
J
Abbasi
A
A
Zaidi
Z H
ZH
Voelter
W
W
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004487:Edema / Q000139:chemically induced; D004546:Elapid Venoms / Q000737:chemistry* / Q000201:enzymology; D005260:Female; D006461:Hemolysis / Q000187:drug effects; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000633:toxicity; D007928:Lethal Dose 50; D008297:Male; D013058:Mass Spectrometry; D051379:Mice; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity; D054467:Phospholipases A2; D051381:Rats; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D004546:Elapid Venoms; D007527:Isoenzymes; D010741:Phospholipases A; D054467:Phospholipases A2