Purification and preliminary characterisation of praelongin phospholipases, antiplatelet agents from the snake venom of Acanthophis praelongus.
Three praelongin phospholipases were chromatographically purified from the snake venom of Acanthophis praelongus. The purity and homogeneity of the praelongins were assessed by RP-HPLC, HPCE and mass spectrometry. The purified enzymes, praelongins 2bIII, 2cII and 2cIV were found to have phospholipase A2 activities with specific activities of 31.4 +/- 0.4, 326.1 +/- 10.2 and 362.5 +/- 12.0 U/mg, respectively. Mass spectrometry studies showed the molecular mass of praelongin 2bIII to be 12,782.9 +/- 2.6 and praelongins 2cII and 2cIV to have very similar molecular mass values, 12,971.4 +/- 4.5 and 12,971.9 +/- 3.6, respectively. However, platelet aggregation studies showed the praelongins to display different IC50 values, 180 microM for praelongin 2cII and 55 microM for praelongin 2cIV; praelongin 2bIII was found to be a more potent antiplatelet agent, having an IC50 of 0.65 microM. Praelongins 2bIII, 2cIV and 2cII were found to have pI values of 10.3 +/- 0.3, 9.6 +/- 0.6 and 9.4 +/- 0.6 as determined by HPCE. The antiplatelet potencies do not correspond to their in vitro phospholipase catalytic potencies, but appear to be related to the enzyme isoelectric points.
Sim
K L
KL
Bioscience Centre, Faculty of Science, National University of Singapore, Singapore. bscsimkl@nus.sg
D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004546:Elapid Venoms / Q000737:chemistry / Q000201:enzymology* / Q000494:pharmacology; D019075:Electrophoresis, Capillary; D006801:Humans; D066298:In Vitro Techniques; D007526:Isoelectric Point; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D013058:Mass Spectrometry; D008970:Molecular Weight; D010741:Phospholipases A / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D054467:Phospholipases A2; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D011517:Prothrombin Time
D004546:Elapid Venoms; D007527:Isoenzymes; D010975:Platelet Aggregation Inhibitors; D010741:Phospholipases A; D054467:Phospholipases A2