PubMed · 562620
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J R Challis, M J Bennett. 1977-11-15. Cortisol binding in human amniotic fluid.. https://doi.org/10.1016/0002-9378(77)90648-2
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Chimeric cowpea mosaic virus (CPMV) particles displaying foreign peptide antigens on the particle surface are suitable for development of peptide-based vaccines. However, commonly used PEG precipitation-based purification methods are not sufficient for production of high quality vaccine candidates because they do not allow for separation of chimeric particles from cleaved contaminating species. Moreover, the purified particles remain infectious to plants. To advance the CPMV technology further, it is necessary to develop efficient and scalable purification strategies and preferably eliminate the infectivity of chimeric viruses. CPMV was engineered to display a 25 amino acid peptide derived from the Bacillus anthracis protective antigen on the surface loop of the large coat protein subunit. The engineered virus was propagated in cowpea plants and assembled into chimeric virus particles displaying 60 copies of the peptide on the surface. An effective inactivation method was developed to produce non-infectious chimeric CPMV virus-like particles (VLPs). Uncleaved VLPs were separated from the contaminating cleaved forms by anion exchange chromatography. The yield of purified chimeric VLPs was 0.3 g kg(-1) of leaf tissue. The results demonstrate the ability to generate multi-gram quantities of non-infectious, chimeric CPMV VLPs in plants for use in the development of peptide-based vaccines.
(14)N-NMR and (31)P-NMR have been used to monitor the in vivo pH in roots, stems, and needles from seedlings of Norway spruce, a typical ammonium-tolerant plant. The vacuolar and cytoplasmic pH measured by (31)P-NMR was found to be c. pH 4.8 and 7.0, respectively, with no significant difference between plants growing with ammonium or nitrate as the N-source. The (1)H-coupled (14) NH 4+ resonance is pH-sensitive: at alkaline pH it is a narrow singlet line and below pH 4 it is an increasing multiplet line with five signals. The pH values in ammonium-containing compartments measured by (14)N-NMR ranged from 3.7 to 3.9, notably lower than the estimated pH values of the P(i) pools. This suggests that, in seedlings of Norway spruce, ammonium is stored in vacuoles with low pH possibly to protect the seedlings against the toxic effects of ammonium ( NH 4+) or ammonia (NH3). It was also found that concentrations of malate were 3-6 times higher in stems than in roots and needles, with nitrate-grown plants containing more malate than plants grown with ammonium.
The global gene expression profile of Bacillus subtilis in response to ammonium and tryptophan starvation was analyzed using transcriptomics and proteomics which gained novel insights into these starvation responses. The results demonstrate that both starvation conditions induce specific, overlapping and general starvation responses. The TnrA regulon, the glutamine synthetase (glnA) as well as the sigma(L)-dependent bkd and roc operons were most strongly and specifically induced after ammonium starvation. These are involved in the uptake and utilization of ammonium and alternative nitrogen sources such as amino acids, gamma-aminobutyrate, nitrate/nitrite, uric acid/urea and oligopeptides. In addition, several carbon catabolite-controlled genes (e.g. acsA, citB), the alpha-acetolactate synthase/-decarboxylase alsSD operon and several aminotransferase genes were specifically induced after ammonium starvation. The induction of sigma(F)- and sigma(E)-dependent sporulation proteins at later time points in ammonium-starved cells was accompanied by an increased sporulation frequency. The specific response to tryptophan starvation includes the TRAP-regulated tryptophan biosynthesis genes, some RelA-dependent genes (e.g. adeC, ald) as well as spo0E. Furthermore, we recognized overlapping responses between ammonium and tryptophan starvation (e.g. dat, maeN) as well as the common induction of the CodY and sigma(H) general starvation regulons and the RelA-dependent stringent response. Many genes encoding proteins of so far unknown functions could be assigned to specifically or commonly induced genes.