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A high content of methionine sulfoxide was observed in the resiliums (internal hingeligaments) of surf clams. As no isolation procedure which might cause the oxidation of methionine to methionine sulfoxide was involved and the hydrolysis was carried out in vacuo, it is the first solid evidence for the presence of methionine sulfoxide as a constituent of natural protein.

The amino groups of Laticauda semifasciata III (LsIII) purified from the venom of a sea snake, Laticauda semifasciata, were acetylated with acetic anhydride. Three monoacetyl derivatives of LsIII, namely, [1-N2-acetyl-arginine]-LsIII, [23-N6-acetyl-lysine]-LsIII and [35-N6-acetyl-lysine]-LsIII, were obtained. These monoacetyl derivatives of LsIII showed the same CD spectra as that of native LsIII. [1-N2-Acetyl-arginine]-LsIII was half as active as the original toxin suggesting the importance of the net positive charge of the toxin molecule for the toxicity. [23-N6-Acetyl-lysine]-LsIII and [35-N6-acetyl-lysine]-LsIII showed no toxicity. Lysine-23 is one of the residues common to all neurotoxins, whereas lysine-35 is one of the residues found specifically among long-chain neurotoxins, although there are some exceptional toxins without lysine-23 or lysine-35.

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.

A "non-divergence theory" is proposed for the mechanism of evolution. The theory is based on the observation that comparison of the amino acid sequences of related proteins in various organisms gives inconsistent results from one type of protein to another, and on the occurrence of significant gene transfer among living organisms. Special attention is focused on the sequence comparisons of short- and long-chain neurotoxins and phospholipases A2 from the venoms of proteroglyphous snakes and those of microbial ferredoxins, rubredoxins, and flavodoxins.

The 270-MHz proton NMR spectra of the unique long neurotoxins bearing Phe-25, Astrotia stokesii b (As b) and Astrotia stokesii c (As c) from Astrotia stokesii, and Acanthophis antarcticus b (Aa b) from Acanthophis antarcticus, have been analyzed. The aromatic proton resonances of Phe-25 in As b and Aa b were assigned on the basis of the nuclear Overhauser effects observed on irradiation of slowly exchanging amide protons. Phe-25 was found to be involved in hydrophobic interactions with Ile/Val-42, Ala-46 and Ile-58 in As b and As c, and with Ala-46 and Val-58 in Aa b. These hydrophobic interactions, instead of the hydrogen bond between Tyr-25 and Glu-42 found in other neurotoxins, appear to be important for maintenance of the biologically active tertiary structure. The pH dependency of the chemical shift and intensity of the Trp-72 N-1 proton resonance of As b indicates that the indole ring is not fully exposed to the solvent and that the extra tail segment of this long neurotoxin interacts with the main part of the molecule.