Biomedical subjects

D J Ellar

Publications and source records attributed to D J Ellar.

130 records · Page 8Linked to original sources

PubMed1966

Fine structure of sporulation in Bacillus cereus grown in a chemically defined medium.

Ellar, D. J. (Syracuse University, Syracuse, N.Y.), and D. G. Lundgren. Fine structure of sporulation in Bacillus cereus grown in a chemically defined medium. J. Bacteriol. 92:1748-1764. 1966.-A study was made of the fine structure of sporulating cells of Bacillus cereus grown in a chemically defined medium. The developmental stages of sporulation occurred in a fairly synchronous manner and were complete by 14 hr. This time period was shortened when spore wall peptide components were added to the medium, but the addition had no effect upon fine structure except to thicken the cell wall. Sporulation could be separated into six morphological stages which generally agreed with those published for other sporulating bacteria. The initiation of the spore (forespore) septum takes the form of an inward folding of the cytoplasmic membrane toward the pole of the cell. The inward folding forms a characteristic Y-shaped membrane structure enclosing an area within which vesicles are found. These vesicles comprise the perisporal mesosome of the cell. The membranes on opposite sides of the cell progress toward the cell center where they fuse to form the double unit membrane of the spore septum. As the proliferation of the spore septum continues, the vesicular areas move towards the pole. The end result is a double forespore membrane which completely encloses a part of the vegetative cell's chromatin. Sporal mesosomes, as well as membrane vesicles, are involved in the proliferation of the forespore. Vesicles are generally bounded by a single unit membrane, whereas in the sporal mesosomes several unit membranes are arranged concentrically. The latter become associated with the segregation of a portion of the nuclear material into the forespore region of the cell.

Bacillus cereus↗

PubMedDate not supplied

Cell targeting of a pore-forming toxin, CytA delta-endotoxin from Bacillus thuringiensis subspecies israelensis, by conjugating CytA with anti-Thy 1 monoclonal antibodies and insulin.

The cytolytic protein toxin CytA was linked to two monoclonal antibodies (mAb) directed against the mouse or the rat Thy 1 antigen. The purified CytA-mAb conjugates were not toxic to either target or nontarget cells. The conjugates did bind specifically to target cells since they agglutinated the target cells but not nontarget cells. When the conjugates were treated with dithiothreitol, the released CytA was toxic to all cells tested. These results suggested that the attachment of CytA to a molecule such as the mAb prevented it from forming a pore. Another conjugate was made by linking CytA to insulin. The purified insulin-CytA conjugate bound to and intoxicated cells bearing a high number of insulin receptors. Furthermore, the conjugate was far less toxic to cells expressing a low number of insulin receptors and not toxic to a known CytA target cell line from Aedes aegypti. However, CytA released from the conjugate by reduction was toxic to all cells tested. These results suggested that the cytotoxicity exhibited by CytA in the conjugate form against cells bearing insulin receptors was mediated through insulin and that, in the conjugate form, CytA no longer shows its broad in vitro cytolytic activity. The difference in toxicity between CytA-mAb conjugates and insulin-CytA conjugate is discussed in relation to size of the ligands, the number, distribution, and mobility of the target molecules, and intracellular trafficking.

Animals↗

PubMedDate not supplied

Structural and functional studies of a synthetic peptide mimicking a proposed membrane inserting region of a Bacillus thuringiensis delta-endotoxin.

In order to study the mechanism of action of Bacillus thuringiensis delta-endotoxins, a synthetic 31-mer peptide corresponding to the sequence of a putative pore-forming segment of the CrylA(c) toxin was characterized structurally and functionally. The peptide maps onto the central helix (alpha 5) of the six-helix bundle of domain I of the crystal structure of the CryIIIA toxin. CD and NMR spectroscopic studies indicated that the peptide exists as an alpha-helix in methanol and a random coil in water. The peptide associated with liposomes at pH 4.7 and formed discrete, characterizable channels in planar lipid bilayers at low pHs. These channels had a conductance value of 60 picosiemens (pS). It is possible that this helix is a component of the transmembrane pore formed by B. thuringiensis delta-endotoxins in vivo.

Amino Acid Sequence↗

PubMedDate not supplied

Channel activity caused by a Bacillus thuringiensis delta-endotoxin preparation depends on the method of activation.

The spontaneous insertion of Bacillus thuringiensis Cry delta-endotoxins into planar lipid bilayers to form discrete channels in the absence of receptors is the subject of conflicting reports in the literature. Because these proteins are synthesized as protoxins requiring proteolytic activation for conversion to the active form, differences in the in-vitro protocol used for this activation could be responsible for the contradictory results. To investigate this, CrylA(c) toxin was activated by different procedures, and its ability to release glucose entrapped within liposomes and to form channels in planar lipid bilayers assessed. The toxin preparations exhibited widely differing activities on the lipid membranes; SDS-PAGE and immunoblot analysis suggested that variations in the protein profile of the activated samples could be responsible. These findings raise important practical considerations for further in-vitro studies into the mechanism of action of these toxins.

Bacillus thuringiensis↗