Hydrothermal fabrication of selectively doped organic assisted advanced ZnO nanomaterial for solar driven photocatalysis.

Hydrothermal fabrication of selectively doped (Ag(+)+Pd(3+)) advanced ZnO nanomaterial has been carried out under mild pressure temperature conditions (autogeneous; 150°C). Gluconic acid has been used as a surface modifier to effectively control the particle size and morphology of these ZnO nanoparticles. The experimental parameters were tuned to achieve optimum conditions for the synthesis of selectively doped ZnO nanomaterials with an experimental duration of 4 hr. These selectively doped ZnO nanoparticles were characterized using powder X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), UV-Vis spectroscopy and scanning electron microscopy (SEM). The solar driven photocatalytic studies have been carried out for organic dyes, i.e., Procion MX-5B dye, Cibacron Brilliant Yellow dye, Indigo Carmine dye, separately and all three mixed, by using gluconic acid modified selectively doped advanced ZnO nanomaterial. The influence of catalyst, its concentration and initial dye concentration resulted in the photocatalytic efficiency of 89% under daylight.

Human parainfluenza virus type 1 phosphoprotein is constitutively phosphorylated at Ser-120 and Ser-184.

RNA-dependent RNA polymerases of single-stranded, negative-sense RNA viruses comprise a phosphoprotein (P) and a large protein. The constitutive phosphorylation of the P protein in these viruses is highly conserved, yet the functional significance of phosphorylation is enigmatic. To approach this problem, phosphorylation sites were determined in two closely related paramyxovirus P proteins. Sendai virus (SV) is a prototypic paramyxovirus. Previously, using a phosphopeptide mapping technique, the primary constitutive phosphorylation site of SV P protein was mapped to Ser-249. Phosphorylation at Ser-249 is dependent on the presence of Pro-250. Human parainfluenza virus type 1 (HPIV-1) P protein has 66% similarity to SV P protein and its predicted secondary structure is highly similar to that of SV P protein. However, there is no obvious conserved phosphorylation site in HPIV-1 P protein. Using the phosphopeptide mapping strategy, the constitutive phosphorylation sites of HPIV-1 P protein were mapped. The HPIV-1 P protein is primarily phosphorylated at Ser-120. Phosphorylation at Ser-120 is dependent on the presence of Pro-121. It also has a minor phosphorylation site at Ser-184. The sequence at Ser-184 does not match any consensus phosphorylation target site for the known kinases. Significantly, the P proteins from both viruses are constitutively and primarily phosphorylated at one serine and the phosphorylation of that serine is dependent on the presence of a proline on its carboxyl side.

Sendai virus P protein is constitutively phosphorylated at serine249: high phosphorylation potential of the P protein.

Previously we showed that the Sendai virus P protein (568 aa) in virus-infected cells and in virions was primarily and constitutively phosphorylated on serine(s) in a single tryptic phosphopeptide TP1. By two-dimensional thin-layer electrophoresis and chromatography analysis of tryptic phosphopeptides of several deletion and point mutants of the P protein, we now show that the sole phosphorylation site in TP1 is serine249. Interestingly, when serine249 was deleted or mutagenized alternate potential serine sites were more heavily phosphorylated. A similar effect was observed when the deletion was very close to serine249 (delta 208-236). Mutagenesis of proline250 to alanine abrogated phosphorylation at serine249 suggesting that proline250 is essential for the primary phosphorylation of the P protein. Conceivably, serine249 phosphorylation is mediated by a proline-directed protein kinase. This finding is unusual because a majority of the P proteins from other negative-strand RNA viruses have been shown to be phosphorylated primarily by casein kinase II. Our results demonstrate that the P protein has a strong potency to remain phosphorylated. Based on our previous and present results, we suggest that the phosphorylation sites on P are dependent on the accessibility of phosphatases rather than kinases as all potential sites are about equally competent for phosphorylation. We propose that phosphorylation is important for maintaining the structural integrity of the Sendai virus P protein.

A highly efficient procedure for site-specific mutagenesis of full-length plasmids using Vent DNA polymerase.

Careful titration of Vent polymerase activity allows efficient amplification of full-length plasmids (12 kb). The high processivity and fidelity of this enzyme made oligonucleotide-directed site-specific mutagenesis of plasmids a straight-forward process. Using only two primers, a mutagenic and a complementary, single-base mutants of recombinant plasmids were obtained consistently with > 90% efficiency from a single round of PCR. This procedure also made site-specific deletion, insertion, and several bases mutagenesis facile and efficient.

Intracellular phosphorylation of the Sendai virus P protein.

Phosphorylation status of the Sendai virus P protein was examined during virus infection and compared with cell-free phosphorylation. P protein from Sendai virus-infected (VI) and P/C gene-transfected (PT) mammalian cells and from purified virions (PV) was phosphorylated at only serine residues. In contrast, cell-free phosphorylation of the P protein with virion-associated protein kinase (VAPK) occurred at both threonine and serine. Tryptic phosphopeptide maps of the P protein from VI, PT, and PV showed that the phosphorylation was primarily localized on one peptide (TP1), while VAPK phosphorylated the P protein on several peptides. There was no change in the steady-state phosphopeptide map of the P protein during virus replication, indicating that the TP1 is constitutively phosphorylated. Inhibition of cellular phosphatases (PP1 and PP2A) by okadaic acid (OA) in virus-infected cells caused a sixfold increase in the P protein phosphorylation, solely at serine residues. The phosphopeptide map of the OA-P protein revealed that phosphorylation occurred on several peptides, but the OA-P map was significantly different from the VAPK-P map. However, additional phosphorylation of the P protein did not block its association with nucleocapsids. These results suggest that the Sendai virus P protein is constitutively phosphorylated primarily at one locus but has the potential for phosphorylation at additional sites. Further, our results do not show any correlations between the intracellular and cell-free phosphorylation of the P protein and, therefore, question the validity of cell-free phosphorylations.