No abstract

A recombinant transmissible gastroenteritis coronavirus (rTGEV) in which E gene was deleted (rTGEV-DeltaE) has been engineered. This deletion mutant only grows in cells expressing E protein (E(+) cells) indicating that E was an essential gene for TGEV replication. Electron microscopy studies of rTGEV-DeltaE infected BHK-pAPN-E(-) cells showed that only immature intracellular virions were assembled. These virions were non-infectious and not secreted to the extracellular medium in BHK-pAPN-E(-) cells. RNA and protein composition analysis by RNase-gold and immunoelectron microscopy showed that rTGEV-DeltaE virions contained RNA and also all the structural TGEV proteins, except the deleted E protein. Nevertheless, full virion maturation was blocked. Studies of the rTGEV-DeltaE subcellular localization by confocal and immunoelectron microscopy in infected E(-) cells showed that in the absence of E protein virus trafficking was arrested in the intermediate compartment. Therefore, the absence of E protein in TGEV resulted in two actions, a blockade of virus trafficking in the membranes of the secretory pathway, and prevention of full virus maturation.

No abstract

Nanoscopic and uniform unilamellar vesicles, rich in cholesterol, have been prepared by a new procedure, named "DELOS-SUSP", which is based on the depressurization of a cholesterol solution in CO 2 -expanded acetone into an aqueous solution containing a surfactant. The CO 2 is used here as a cosolvent medium, allowing the straightforward preparation of vesicular systems with controlled size distribution, uniform shape, and stability unachievable by conventional mixing technologies. The resulting nanoscopic vesicular systems dispersed in water were characterized using dynamic light scattering, ζ potential, turbidity, and cryogenic transmission electron microscopy. The influence of operational parameters of this new methodology on the physicochemical characteristics of the vesicular systems is also reported.