Highly polymorphic vitelline-coat protein HaVC80 from the ascidian, Halocynthia aurantium: Structural analysis and involvement in self/nonself recognition during fertilization

While all ascidians (Urochordata) are hermaphroditic, some, including Halocynthia roretzi (Fuke, 1983) and Ciona intestinalis (Rosati & De Santis, 1978) are strictly self-sterile because of a self–nonself recognition system in the interaction between the sperm and the vitelline coat of the eggs. However, immature oocytes (Fuke & Numakunai, 1996) and acidic-seawater-treated mature oocytes (Morgan, 1939; Kawamura et al., 1991) are self-fertile. It is thought that a putative self–nonself recognition molecule, which is detached or modified by treatment with acidic seawater, may be attached to the vitelline coat during oocyte maturation. Although the existence of a self–nonself recognition system in the fertilisation process is well known, the molecular entity has yet to be conclusively identified. However, there have been several attempts to identify such a molecule in Ciona (Marino et al., 1999). In the present study, we have isolated and analysed a molecule which appears to be responsible for allorecognition in the interaction between sperm and eggs of the ascidian Halocynthia roretzi.Biologicals. A solitary ascidian Halocynthia roretzi Type C was used in this study. The fertilisation experiment was carried out as described previously (Sawada et al., 1982).Isolation and N-terminal Sequencing of Hr VC70. Vitelline coats were isolated from immature and mature oocytes of the ascidian by homogenisation and repeated washing with 5× diluted artificial seawater. The isolated vitelline coats were subjected to SDS-PAGE, followed by blotting to a PVDF membrane. The N-terminal amino acid sequence of the 70 kDa main component (HrVC70) was determined by a protein sequencer.

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Structural analysis of the surface-layer protein of spirillum serpens by high-resolution electron microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100077268 ◽

1983 ◽

Vol 41 ◽

pp. 738-739

Author(s):

W. H. Wu ◽

R. M. Glaeser

Keyword(s):

Electron Microscopy ◽

Surface Layer ◽

Structural Analysis ◽

High Resolution ◽

Low Dose ◽

High Resolution Electron Microscopy ◽

Optical Diffraction ◽

Surface Layer Protein ◽

Morphological Unit ◽

Resolution Electron

Spirillum serpens possesses a surface layer protein which exhibits a regular hexagonal packing of the morphological subunits. A morphological model of the structure of the protein has been proposed at a resolution of about 25 Å, in which the morphological unit might be described as having the appearance of a flared-out, hollow cylinder with six ÅspokesÅ at the flared end. In order to understand the detailed association of the macromolecules, it is necessary to do a high resolution structural analysis. Large, single layered arrays of the surface layer protein have been obtained for this purpose by means of extensive heating in high CaCl2, a procedure derived from that of Buckmire and Murray. Low dose, low temperature electron microscopy has been applied to the large arrays.As a first step, the samples were negatively stained with neutralized phosphotungstic acid, and the specimens were imaged at 40,000 magnification by use of a high resolution cold stage on a JE0L 100B. Low dose images were recorded with exposures of 7-9 electrons/Å2. The micrographs obtained (Fig. 1) were examined by use of optical diffraction (Fig. 2) to tell what areas were especially well ordered.

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RNA polymerase: Preparation and structural analysis of helical polymers

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010011091x ◽

1984 ◽

Vol 42 ◽

pp. 152-153

Author(s):

E. Loren Buhle ◽

Pamela Rew ◽

Ueli Aebi

Keyword(s):

Structural Analysis ◽

Rna Polymerase ◽

Molecular Level ◽

X Ray Diffraction ◽

Helical Polymers ◽

X Ray ◽

E Coli ◽

The Past ◽

Ordered Arrays ◽

Low Ionic Strength

While DNA-dependent RNA polymerase represents one of the key enzymes involved in transcription and ultimately in gene expression in procaryotic and eucaryotic cells, little progress has been made towards elucidation of its 3-D structure at the molecular level over the past few years. This is mainly because to date no 3-D crystals suitable for X-ray diffraction analysis have been obtained with this rather large (MW ~500 kd) multi-subunit (α2ββ'ζ). As an alternative, we have been trying to form ordered arrays of RNA polymerase from E. coli suitable for structural analysis in the electron microscope combined with image processing. Here we report about helical polymers induced from holoenzyme (α2ββ'ζ) at low ionic strength with 5-7 mM MnCl2 (see Fig. 1a). The presence of the ζ-subunit (MW 86 kd) is required to form these polymers, since the core enzyme (α2ββ') does fail to assemble into such structures under these conditions.

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Interpreting HVEM of muscle-cell impulse networks

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010012103x ◽

1992 ◽

Vol 50 (1) ◽

pp. 126-127

Author(s):

Paul DeCosta ◽

Kyugon Cho ◽

Stephen Shemlon ◽

Heesung Jun ◽

Stanley M. Dunn

Keyword(s):

Structural Analysis ◽

Muscle Cell ◽

Electron Micrographs ◽

Relative Size ◽

Automatic Classification ◽

Nerve Fibers ◽

Analysis Algorithm ◽

Structural Networks

Introduction: The analysis and interpretation of electron micrographs of cells and tissues, often requires the accurate extraction of structural networks, which either provide immediate 2D or 3D information, or from which the desired information can be inferred. The images of these structures contain lines and/or curves whose orientation, lengths, and intersections characterize the overall network.Some examples exist of studies that have been done in the analysis of networks of natural structures. In, Sebok and Roemer determine the complexity of nerve structures in an EM formed slide. Here the number of nodes that exist in the image describes how dense nerve fibers are in a particular region of the skin. Hildith proposes a network structural analysis algorithm for the automatic classification of chromosome spreads (type, relative size and orientation).

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