Odile Sorokine

PURIFICATION AND CHARACTERIZATION OF ANDROGENIC HORMONE FROM THE TERRESTRIAL ISOPOD ARMADILLIDIUM VULGARE LATR. (CRUSTACEA, ONISCIDEA)

Androgenic hormone (AH) was purified from hypertrophied androgenic glands of intersexed Armadillidium vulgare (genetic males feminized by symbiotic endocellular bacteria). Two isohormones labeled AH1 and AH2 with similar molecular weights in the range 17,000-18,000 were isolated. Amino acid analysis showed the absence of cysteine in these two forms. A polyclonal antiserum was raised which recognized AH1 and AH2. The physiological significance of this polymorphism is still not known.

Developmental Study of Proteolipids in Bovine Brain: A Novel Proteolipid and DM-20 Appear Before Proteolipid Protein (PLP) During Myelination

Abstract: In a developmental study, we have shown that DM‐20 is present before proteolipid protein (PLP) in the fetal bovine cerebral hemispheres. When the white matter appears (27–30 weeks of gestation), the amount of DM‐20 drastically increases. DM‐20 remains the major proteolipid until birth. PLP is detected only 2–4 weeks after the appearance of white matter, that is, more than 4 weeks after the appearance of DM‐20. The early appearance of DM‐20 at the beginning of myelination raises the question of its particular function. In the adult bovine cerebral hemispheres, PLP is the major proteolipid but DM‐20 remains quantitatively important because the PLP/DM‐20 ratio ranges from 1.5 to 1.7. In the same developmental study we have, in the fetal cerebral hemispheres, isolated and characterized a novel proteolipid (apparent Mr 20,000), which appears even before DM‐20 and is not detected in the adult brain. It is structurally related to PLP and DM‐20 because the first 31 N‐terminal amino acid residues are the same. However, in immunoblot. it did not react either with the antitridecapeptide 117–129 antiserum of PLP or with the anti‐C‐terminal hexapeptide antiserum of PLP.

Aah VI, a novel, N-glycosylated anti-insect toxin from Androctonus australis hector scorpion venom: isolation, characterisation, and glycan structure determination

Aah VI was isolated from the venom of the North African scorpion, Androctonus australis hector. It is the first glycosylated neurotoxin from scorpion venom to be described. It was not toxic to mice, when injected intracerebroventricularly at a dose of 1.2 μg per animal. However, it had typical activity in Blatella germanica cockroaches resulting in gradual paralysis and very low toxicity (LD50=8.5 μg/g of animal). It consists of 66 amino acid residues and is heterogeneously N‐glycosylated at a single site, on asparagine 9, of the Asn‐Gly‐Thr sequence. The potential N‐glycosylation site was deduced from automatic Edman degradation and amino acid analysis, and glycan heterogeneity was evidenced by ESMS. Determination of the N‐glycan structures (dHex, Hex and HexNAc) was assessed by nanoESMS/MS with picomolar amounts of sample. Current knowledge of N‐glycan structure and composition suggests that the glycan structures are derived from a common core.

Purification and characterization of minor brain proteolipids: use of fast atom bombardment — mass spectrometry for peptide sequencing

A combination of lipophilic gel permeation chromatography and ion-exchange chromatography in organic solvents was used to purify low molecular weight proteolipids from bovine brain. Cleavage peptides were purified by HPLC and studied mainly by the fast atom bombardment--mass spectrometry technique. A proteolipid of Mr 14 000 contains several peptides from the first 113 amino acids of the major myelin proteolipid (MMPL) plus an extra unknown blocked N-terminal peptide. A proteolipid of Mr 16 000 contains smaller peptides belonging to a C-terminal fragment of MMPL of about 160 residues. These two proteolipids do not seem to be artifacts from MMPL.

Specific proteolytic cleavage of the myristoylated alanine‐rich C kinase substrate between Asn 147 and Glu 148 also occurs in brain

The myristoylated alanine‐rich C kinase substrate (MARCKS) is a major ubiquitous substrate of protein kinase C. The expression of the protein is regulated during cell cycle progression and cell proliferation. Specific proteolytic cleavage of the protein between Asn 147 and Glu 148 was described recently in cultured cells, and the corresponding proteolytic activity was observed in various tissue extracts except for brain. We purified a 40 kDa fragment of MARCKS from bovine brain that we characterized as the C‐terminal specific fragment found in other tissues. The identification of the fragment was achieved by in vitro phosphorylation by protein kinase C, calcium‐dependent interaction with calmodulin, mass spectrometric analysis, and N‐terminal sequencing. These data suggest that specific proteolytic cleavage of MARCKS also occurs in brain and may be a general mechanism of down‐regulation of the protein. J. Neurosci. Res. 48:259–263, 1997.

Marcks, a Major in Vivo Substrate of Protein Kinase C Purification, Interaction with Model Membrane, and Demyristoylation

The protein kinase C (PKC) family has been implicated in a large number of cellular responses. In order to better understand the cellular function of these enzymes, the identification of physiologically important substrates has received a peculiar attention during the last decade. The myristoylated alanine-rich C kinase substrate (MARCKS) is a widely distributed in vivo and in vitro substrate that has been often used as a marker of the activation of the kinase. (see Aderem, 1992 and Blackshear, 1993). Its phosphorylation by PKC in the cells leads to its translocation from the plasma membrane to the cytoplasmic compartment. The process is reversible, the dephosphorylation by cellular phosphatases leading to its reassociation with the plasma membrane (Thelen et al., 1991).