Phospholipase A of Sea Snake Laticauda semjfasciata Venom

Abstract

The venom gland extracts of the sea snake Laticauda semifasciata contained at least four forms of phospholipase A separable on a CM-cellulose column. They were designated as phospholipases A I-IV in the order of elution from the column. Phospholipases A I, III, and IV were isolated in a homogeneous state. They were similar to one another in amino acid composition and molecular weight (14,000) as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Phospholipase A I contained one tryptophan residue. whereas III and IV did not. Although all these forms had the same A2-type positional specificity, they were classified into two groups (I, and III and IV) on the basis of enzymic properties. Phospholipase A I had a higher specific activity and showed normal kinetics, whereas III and IV had approximately one-tenth of the specific activity of I and showed biphasic kinetics due to their activation by the reaction products. Phospholipase A I, the major form, seems to be identical with phospholipase A reported previously (Tu, A.T., Passey, R.B., & Toom, P.M. (1970) Arch. Biochem. Biophys. 140, 96-106), whereas the other two, III and IV, are new. Phospholipase A I became more like III and IV in enzymic properties on modification with N-bromosuccinimide.

Isolation, properties and amino acid sequences of a phospholipase A2 and its homologue without activity from the venom of a sea snake, Laticauda colubrina, from the Solomon Islands

“…LcPLA-II showed biphasic kinetics due to activation by the reaction product (oleate). A similar observation was reported on the L. semifasciata phospholipases A2 LsPLA III and LsPLA IV (Yoshida et al, 1979). The addition of oleate shortened the lag period without changing the highest velocity, similarly to the cases of LsPLA III and LsPLA IV.…”

“…LsPLA I, which shows normal kinetics, can be converted into LsPLA III or LsPLA IV type with biphasic kinetics on the oxidation of Trp-64 (Yoshida et al, 1979). LcPLA-II has Leu-64, similarly to LsPLA III or LsPLA IV.…”

“…The venom was milked from the snakes, diluted with an equal volume of 0.2 M-acetic acid, brought back to the laboratory and freeze-dried. Materials Phospholipases A2 LsPLA I, LsPLA III and LsPLA IV from Laticauda semifasciata venom were purified as described previously (Yoshida et al, 1979). Radioactive 45CaC12 (specified radioactivity 1.06 Ci/mmol) was obtained from Amersham International (Amersham, M, determination Mr determination by gel filtration was carried out with the Pharmacia f.p.l.c.…”

“…Ca21 was required and the maximal activity was attained with 3 mM-CaCl2 in the presence of 0.7 mM-EDTA. The specific activity of the preparation from fraction B-1I measured by a titrametric method (Yoshida et al, 1979) was 24 ,mol/min per mg with egg-yolk phosphatidylcholine (4 mM) as a substrate at 37°C in the presence of 8 mM-Triton X-100, 3mM-CaCl2, 0.7 mM-EDTA and 1.4 mM-sodium oleate at pH 8.2 (Table 2). On the other hand, the preparation from fraction B-I showed no phospholipase A2 activity in the presence or in the absence of 1.4 mM-sodium oleate and in the presence of 3 mm-or 30 mM-CaCl2.…”

“…Furthermore, a non-neurotoxic non-enzymic phospholipase A2 homologue, Notechis II-1, was isolated from an Australian elapid snake, Notechis scutatus scutatus , and its amino acid sequence determined (Lind & Eaker, 1980). From a member of the sea kraits, Laticauda semifasciata, three non-neurotoxic phospholipases A2, LsPLA I, LsPLA III and LsPLA IV, were isolated (Yoshida et al, 1979) and the amino acid sequences determined (Nishida et al, 1982).…”