The mRNA encoding human thyroglobulin has been cloned and sequenced. It is made up of a 8301-nucleotide segment encoding a preprotein monomer of 2767 amino acids, flanked by non-coding 5' and 3' regions of 41 and 106 nucleotides, respectively. This preprotein consists of a leader sequence of 19 amino acids, followed by the sequence of the mature monomer, corresponding to a polypeptide of 2748 amono acids ( M , = 302 773). On its amino-terminal side, 70% of the monomer is characterized by the presence of three types of repetitive units. In contrast, the remaining 30% of the protein is devoid of repetitive units. This last region however shows an interesting homology (up to 64%) with the acetylcholinesterase of Torpedo californica. The sites of thyroid hormones synthesis are clustered at both ends of the thyroglobulin monomer. By contrast, the potential glycosylation sites are scattered along the polypeptide chain.Thyroglobulin is a protein specifically synthesised by the thyroid gland, and constitutes the support for the production of the two thyroid hormones, thyroxine and triiodothyronine [l]. The existence of thyroglobulin was demonstrated a century ago [2], but its structure has been elucidated only recently. It is a dimeric glycoprotein with an M , of 660000, of two identical subunits [3, 41 encoded by a single mRNA with a sedimentation coefficient of 33 S (8500 nucleotides) [5 -71. Thyroglobulin is synthesised by the thyrocyte, then exported to the vesicular lumen where its maturation begins by the iodination of several tyrosine residues, and coupling of some of the iodotyrosine residues [8]. Then, by an endocytotic process, the molecule is absorbed into the thyrocyte where several selective cleavages occur in the lysosomes, resulting in the release of the thyroid hormones, and complete degradation of the rest of the molecule.For a thyroglobulin iodine content of 0.5% (which is rarely attained in man) a maximum of 3.5 hormonal residues per thyroglobulin molecule are formed through a reaction catalyzed by the enzyme thyroid peroxidase [9]. Four hormone-synthesis sites have been described, corresponding to four tyrosine residues in fixed positions [lo-121.The structure of human thyroglobulin seems to be responsible for the specific fixation of iodine, and the production of functional thyroid hormones. Several human pathologies are associated with an abnormal thyroid function. Since the recent demonstration of the implication of a defect in thyroglobulin gene structure in the development of congenital goitre in cattle [13], it is likely that knowledge of the structure of human thyroglobulin mRNA will help to elucidate the structural bases of human thyroid pathologies. We describe here the complete nucleotide sequence of human thyroglobulin mRNA. MATERIALS AND METHODSPreparation and sequencing of DNA cDNA fragments corresponding to human thyroglobulin mRNA were prepared from recombinant plasmids named M1-M4 and B2 -B4 (see Fig. l), as previously described [7, 141. Two additional clones, named B1 (kind gift of ...

Human thyroid peroxidase (TPO) has been purified from thyroid microsomes by immunoaffrnity chromatography using a monoclonal antibody (mAb) to TPO. The eluted material had a specific activity of 381 U/mg and exhibited a peak in the Soret region. The ratio of A,,, to A,,, ranged from 0.20 to 0.25. Upon SDS-polyacrylamide gel electrophoresis, the purified enzyme gave two contiguous bands in the 100 kDa region. Further, it has been demonstrated that sera with anti-microsomal autoantibodies from patients presenting Graves' or Hashimoto's thyroiditis diseases were able to bind to purified TPO and to inhibit in a dose-dependent manner the mAb binding to purified TPO. This suggests that TPO is the thyroid antigen termed to date the microsomal antigen. Microsomal antigen Thyroidperoxidase AutoantibodyMonoclonal antibody Autoimmunity

The purification of eleven neurotoxins from the venom of three scorpions (Androctonus australis Hector, Buthus occitanus tunetanus and Leiurus quinquestriatus quinquestriatw8) has been performed by Sephadex G-50 filtration (with and without recycling) and equilibrium chromatography on Amberlite CG-50, CM-Sephadex C-50 and DEAE-Sephadex A-50. All the chromatographic steps were carried out in ammonium acetate buffers. Purity of the neurotoxins was assessed by amino acid analysis and zone electrophoresis. All have a molecular weight of about 7000 daltons and do not contain methionine. As shown by amino acid composition and chain length scorpion neurotoxins show a high degree of homology.From the first attempts made in this laboratory to purify the neurotoxins of scorpion venoms, successive improvements in the purification method have been introduced. Snake or scorpion neurotoxins are basic proteins of small molecular weight (about 7000). Two properties of these neurotoxins, solubility in pure water [l] and basicity as shown by zone electrophoresis [2], served as guide-lines to use reversible adsorption on Sephadex G-25 swollen in water [3] and ion-exchange chromatography on Amberlite IRC-50 in a volatile buffer a t near neutral pH, to obtain the purified neurotoxins [2,4,5]. Despite homogeneity in the ultracentrifuge, by starch gel electrophoresis and by equilibrium chromatography on Amberlite IRC-50, further studies have shown the purified proteins to be dimers of non-covalently associated monomers or non-covalent association of neurotoxins with non-toxic material Two pure monomeric neurotoxins from the NorthAfrican scorpion Androctonus australis Hector collected in the area of Tozeur (Tunisia) [ill were later obtained with Sephadex G-50 filtration, chromatography on Amberlite CG-50 a t pH 6.30 and 6.70 and on DEAE-Sephadex A-50 a t alkaline p H (8.50) still in volatile buffer.This and the following paper describe a general method of purification of the neurotoxins contained in animal venoms (scorpions and snakes). Improvement over the previously described method [ll] was obtained by introducing recycling gel filtration on Sephadex and the use of another cation-exchanger, CM-Sephadex C-50. I n this first paper we describe the isolation and the characterization of eleven pure EXPERIMENTACPROCEDURE MATERIALS VenomsThe venoms of Buthus occitanw tunetanus (Mecheria, Algeria) and Leiurus quinquestriatus quinquestriatus (area of Khartum, Sudan) were obtained from F. G . Celo (Zweibriicken, Germany). The scorpions Androctonus australis Hector were collected in the area of Chellala (Algeria) by the Institut Pasteur of Algeria. They were maintained alive in our laboratory in individual plastic boxes and received food (mealworms) and water via soaked sponges each month. The animals were kept a t 22-25" except for the week preceding milking during which the temperature was raised to 30-33". Milking was obtained by cyclic electrical stimulations of the postabdomen (55 volts for 0.5 sec followed by a rest of the same duratio...