Leukemogenic activity of ether-extracted Rauscher leukemia virus.

After rapid multiple extractions of mouse plasma virus with ether, the aqueous solution contained viral nucleoids that were infectious when inoculated intracranially into newborn BALB/c mice. The infectivity associated with the ether extract was not neutralized by the specific antibody prepared against the whole virus. No intact virus has been seen in these preparations. Treatment with ether completely removed the virus envelope from the particle and produced an apparently homogeneous preparation of viral nucleoids. After the extractions with ether, leukemogenic activity was inactivated by exposure to ribonuclease. The leukemogenic activity of the many-passaged Rauscher virus that has been propagated in tissue culture and that has low infectivity was also retained, and, in two experiments in which material was inoculated intracranially into mice, this activity appeared to have been enhanced by multiple extractions with ether.

Bacteriorhodopsin crystal growth in reduced gravity--results under the conditions, given in CPCF on board of a Space Shuttle, versus the conditions, given in DCAM on board of the Space Station Mir.

For the purpose of bio-electronics, bacteriorhodopsin was crystallized into two habits through liquid-liquid-diffusion, namely individual needles of up to 1.9 mm in length and needle bunch-like clusters of up 4.9 mm in total length. In both the reduced gravity experiments performed, the morphology of the individual needles (crystal form A) had improved in terms of sharp needle edges and compact needle packing, compared to the parallel ground controls. For the long duration wide range low gravity condition in the "Diffusion-controlled Crystallization Apparatus for Microgravity (DCAM)" on Mir (STS-89 up), needle bunches on average were longer there than on the ground, while the compactness of the clusters, i.e. the average ratio of clustered length to clustered width was the reverse. Some exceptionally large individuals needles were grown in DCAM. For the "Commercial Protein Crystallization Facility (CPCF)" in short duration high definition microgravity conditions during a science mission of the Space Shuttle Discovery (STS-95), size and shape of the individual needles were homogeneous and superior to those of both the parallel ground controls and the results in DCAM. In CPCF, the average volume of the individual needles in suspension was increased by 50% in microgravity compared to those in the ground control.

Immunoprecipitation of the parathyroid hormone receptor.

An 125I-labeled synthetic analog of bovine parathyroid hormone, [8-norleucine,18-norleucine,34-tyrosine]PTH-(1-34) amide ([Nle]PTH-(1-34)-NH2), purified by high-pressure liquid chromatography (HPLC), was employed to label the parathyroid hormone (PTH) receptor in cell lines derived from PTH target tissues: the ROS 17/2.8 rat osteosarcoma of bone and the CV1 and COS monkey kidney lines. After incubation of the radioligand with intact cultured cells, the hormone was covalently attached to receptors by using either a photoaffinity technique or chemical (affinity) cross-linking. In each case, covalent labeling was specific, as evidenced by a reduction of labeling when excess competing nonradioactive ligand was present. After covalent attachment of radioligand, membranes were prepared from the cells and solubilized in the nonionic detergent Nonidet P-40 or octyl glucoside. The soluble membrane fraction present in the supernatant of a 100,000 X g centrifugation was incubated with IgG prepared from anti-PTH antiserum generated to the amino-terminal region, residues 1-34, of PTH. The IgG-PTH-receptor complex was precipitated with staphylococcal protein A-Sepharose. Analysis of the immunoprecipitate on NaDod-SO4/polyacrylamide gel electrophoresis followed by autoradiography revealed the presence of a doublet of apparent molecular mass 69-70 kDa. Specifically labeled bands of approximate molecular mass 95 and 28 kDa were also observed. The anti-PTH IgG was affinity purified by passage over a PTH-Sepharose column and used to make an immunoaffinity column. The 70- and 28-kDa bands were also observed after labeled solubilized membrane preparations were allowed to bind to this column and then were eluted by using a [Nle]PTH-(1-34)-NH2-containing buffer or acetic acid. These studies suggest that the use of an anti-PTH antiserum that binds receptor-bound hormone is likely to be a useful step in the further physiochemical characterization and purification of the PTH receptor.