Multiplicity of acidic subunit isoforms of crotoxin, the phospholipase A2 neurotoxin from Crotalus durissus terrificus venom, results from posttranslational modifications

Crotoxin, the major toxin of the venom of the South American rattlesnake, Crotalus durissus terrificus, is made of two subunits: component B, a basic and weakly toxic phospholipase A2, and component A, an acidic and nontoxic protein that enhances the lethal potency of component B. Crotoxin is a mixture of isoforms that results from the association of several isoforms of its two subunits. In the present investigation, we have purified four component A isoforms that, when associated with the same purified component B isoform, produced different crotoxin isoforms, all having the same specific enzymatic activity and the same lethal potency. We further determined by Edman degradation the polypeptide sequences of these four component A isoforms. They are made of three disulfide-linked polypeptide chains (alpha, beta, and gamma) that correspond to three different regions of a phospholipase A2 precursor. We observed that the polypeptide sequences of the various component A isoforms all agree with the sequence of an unique precursor. The differences between the isoforms result first by differences in the length of the various chains alpha and beta, indicating that component A isoforms are generated from the proteolytic cleavage of the component A precursor at very close sites, possibly by the combined actions of endopeptidases and exopeptidases, and second by the possible cyclization of the alpha-NH2 of the N-terminal glutamine residue of chains beta and gamma. These observations indicate that the component A isoforms are the consequence of different posttranslational events occurring on an unique precursor, rather than the expression of different genes.     

Determination of the major zinc fractions in human serum by ultrafiltration

In this paper we report a method for measuring ultrafiltrable zinc in human serum by electrothermal atomic absorption spectrophtometry. We show also that ultrafiltration permits to determine alpha-2 macroglobulin bound zinc and losely bound zinc if a strong zinc ligand (EDTA) is added to serum before ultrafiltration. This last fraction, after deduction of ultrafiltrable zinc, represents roughly all albumin bound zinc. In 20 controls we found that ultrafiltrable zinc amounted 0.311 μmol/L (S.D.=0.117 μmol/L), alpha-2 macroglobulin bound zinc 3.08 μmol/L (S.D.=0.221 μmol/L), and albumin bound zinc 12.11 μmol/L (S.D.=1.95 μmol/L). Our method needs only a small volume of serum, it is simple and rapid but also very accurate and reliable. The losely bound fraction is very dynamic and, representing the physiologically active part of serum zinc, it could be a good marker of zinc deficiency.     

NCα-gem-dimethylated peptoid side chains: A novel approach for structural control and peptide sequence mimetics

The design of linear peptoid oligomers adopting well-defined secondary structures while mimicking defined peptide primary sequences is a major challenge in the context of drug discovery. To this end, chemists have developed cis-inducing peptoid side chains to build robust polyproline type I helices. However, the number of efficient examples remains scarce and chemical diversity accessible through the use of these side chains is limited. Herein, we introduce an array of NCα-gem-dimethylated peptoid residues mimicking proteinogenic amino acids. Submonomer synthesis and block-coupling approaches were explored to access heterooligomers incorporating these novel types of side chains. NMR studies of monomer and trimer models showed that the NCα-gem-dimethylated groups exert complete cis control on the backbone amide conformation. Lastly, a preliminary molecular modeling study gave an insight into the preferred orientation of the substituents of the NCα-gem-dimethyl side chains relative to the peptoid backbone.     

Isolation of Human Clq as a by-Product of Routine Fractionation

Clq can be isolated as a by-product of large scale fractionation. The euglobulins of fraction III are submitted to chromatography on CM-cellulose in a batch-procedure. The active fraction being frequently contaminated by lysozyme the protein can be further purified by gel-filtration in the presence of carbohydrates. The purified Clq is suitable for the detection of immune complexes.