Jean-Christophe Taveau

Topological mapping of 13 epitopes on a subunit of Androctonus australis hemocyanin.

A topological localization of epitopes on the surface of the Aa6 subunit of Androctonus australis hemocyanin has been carried out. First, immunocomplex strings composed of native hemocyanin and monoclonal antibodies were examined in the electron microscope and submitted to an image processing by correspondence analysis. The average images were then compared to a three-dimensional model of the 24-mer suggesting that 11 of the 13 epitopes are located in three zones of the subunit surface. Second, the overlaps between the epitopes were then studied by polyacrylamide gel electrophoresis, competitive binding inhibition, and immunoelectron microscopy. Four groups of epitopes were identified. One group was capable of binding exclusively to the free subunit. The other three groups were identical to those found in immunoelectron microscopy. The data are consistent with the existence of a small number of immunodominant regions on the surface of the Aa6 subunit.

An approach to the architecture of Scutigera coleoptrata hemocyanin by electron microscopy and image processing

Summary— The 6 × 6meric hemocyanin of the centipede Scutigera coleoptrata, prepared by the single layer negative staining technique with uranyl acetate, has been investigated under the electron microscope. Isolated molecules were windowed from selected digitized micrographs. After alignment, they were submitted to correspondence analysis and hierarchical ascendant classification. Two main clusters of molecules were differentiated, and their average images were obtained. A 3D‐model constructed from these images is proposed.

Revisiting the structure of Alvinella pompejana hemoglobin at 20 Å resolution by cryoelectron microscopy

The hemoglobin of the polychaete worm Alvinella pompejana was reconstructed at 20A resolution from frozen-hydrated samples observed by electron microscopy according to the random conical tilt series method. This three-dimensional reconstruction was mirror-inverted with respect to a previous volume published by de Haas et al. in 1996. In order to explain this handedness discrepancy, various 3D reconstructions using different reference volumes were carried out showing that the choice of the first volume was the keystone during the refinement process. The 3D reconstruction volume of A. pompejana Hb presented structural features characteristic of annelid Hbs with two hexagonal layers each comprising six hollow globular subassemblies and a complex of non-heme linker chains. Moreover, the eclipsed conformation of the two hexagonal layers and a HGS architecture similar to that described for Arenicola marina Hb led to the conclusion that A. pompejana Hb belonged to the architectural type II according to the definition of Jouan et al. (2001). A comparison between this cryo-EM volume and X-ray crystallography density maps of Lumbricus terrestris type-I Hb (Royer et al., 2000) showed that the triple stranded coiled coil structures of linker chains were different. Based on this observation, a model was proposed to explain the eclipsed conformation of the two hexagonal layers of type-II Hbs.

An approach to the architecture of hemocyanin by electron microscopy and image processing

The 6 × 6meric hemocyanin of the centipede Scutigera coleoptrata, prepared by the single layer negative staining technique with uranyl acetate, has been investigated under the electron microscope. Isolated molecules were windowed from selected digitized micrographs. After alignment, they were submitted to correspondence analysis and hierarchical ascendant classification. Two main clusters of molecules were differentiated, and their average images were obtained. A 3D-model constructed from these images is proposed.

The Architecture of 4x6-meric Arachnid Hemocyanin

Arthropod Hcs have highly heterogeneous Mws ranging from approximately 450 to 3,600 kDa. For example, lx6-mers of 75 kDa polypeptide chains have been found in crustaceans, 2x6-mers in crustaceans and spiders, 4x6-mers in most chelicerate groups and in the thalassinid shrimps, 6x6-mers in centipedes, and 8x6-mers in merostoms.

Molecular immunoelectron microscopy and image processing: An approach to epitope mapping on high molecular weight proteins

ULTRASTRUCTURE OF ACCESSORY ADRENOCORTICAL NODULES IN WlSTAR RATS. Anna S. BELLONI ( 1 ) , Francesco MUSAJO (2) , Giuseppina MAZZOCCHI (1) and Gastone G. NUSSDORFER ( 1 ) . (1) Department o f Anatomy and (2) 2nd S u r g i c a l C l i n i c , U n i v e r s i t y o f Padua, I t a l y . More than 70% o f b i l a t e r a l l y a d r e n a l e c t o m i z e d Wis t a r r a t s s u r v i v e and w i t h i n t h r e e months deve lop one o r two consp icuous a d r e n o c o r t i c a l nodules (2-3 mm in d i a m e t e r ) , which, thouEh d i s p l a y i n g an obvious histoloEical zonation, are not associated with chromaffin tissue. Middle and inner cells of the nodules are similar to zona fasciculata (ZF) and zona reticularis elements of the adult rat adrenal Eland, res pectively. Middle cells show numerous round mitochondria with vesicular cristae, while inner cells con tain ovoid mitochondria with tubule-convolute cristae. Both cell types possess abundant smooth endopla smic reticulum and few lipid droplets, as well as a very elaborated microvillous system. Conversely, outer (subcapsular) cells of the accessory nodules do not resemble lena Elomerulosa (ZG) elements, since they display ovoid mitochondria with vesicular cristae instead of elonEated orEanelles with tubu lo-laminar cristae. Moreover, morphometry shows that their volume is about two-fold that of the true ZG cells (1400 um3 versus 700 um3). These morpholoEical data accord well with the fact that basal plasm8 concentration of aldosterone, at variance with that of corticosterone, remains very low three months after bilateral adrenalectomy. The lack of differentiated ZG cells in the accessory adrenocortical nod ules could be explained by the very elevated blood level of ACTH (about 5-times hiEher than in shamoperated rats), in fact, chronic ACTH hypersecretion is known to transform ZG cells into ZF elements (i). Some lines of evidence suEEest the existence of a paracrine control of adrenal zona corticalis by zona medullaris (i). We think that accessory adrenocortical nodules, lackinE zona medullaris, could be a Eood experimental model to Eain insiEht into this problem.