New indanone compounds from Onychium japonicum

A new indanone glucoside pteroside M has been isolated from fronds of Onychium japonicumPteridaceae. The structure of its aglycone pterosin M has been established by ¹³C NMR, PMR spectra and degradation with nitric acid.     

Conversion of L-sorbose to L-sorbosone by Gluconobacter melanogenus

Growing cultures, washed cells, and cell-free extracts of Gluconobacter melanogenus IFO 3293 were found to convert L -sorbose to L -sorbosone. The product was identified by thin layer chromatography of the 2, 4-dinitrophenylhydrazone, and by paper partition chromatography using chemically prepared materials as standards. Factors influencing the conversion included incubation temperature and composition of the growth medium. Addition of betaine or choline to the growing cultures stimulated conversion of L -sorbose to L -sorbosone.     

The effects of calcium channel blocking drugs on the thermotropic behavior of dimyristoylphosphatidylcholine

The effects of four calcium channel blocking drugs, diltiazem, verapamil, nimodipine and nisoldipine, on the main phase transition of dimyristoylphosphatidylcholine have been studied by high resolution differential scanning calorimetry. In all cases, the phase transition temperature is lowered, though much more effectively by nimodipine and nisoldipine than by the other two drugs. Nimodipine and nisoldipine markedly reduce the enthalpy of transition while diltiazem and verapamil have no significant effect on the enthalpy within the drug concentration range studied. Analysis of the data in terms of ideal solution theory is presented. X-ray and neutron scattering studies indicate that nimodipine and verapamil differ significantly with respect to their location within a lipid bilayer, and this difference suggests a partial rationalization of the experimental results presented here.     

Regional distribution of immunoreactive endothelin in porcine tissue: abundance in inner medulla of kidney

A specific and sensitive radioimmunoassay for endothelin has been developed. Half maximal inhibition of binding of radioiodinated endothelin was observed at 37 pg/tube and endothelin was detectable as low as 1 pg/tube. With this assay, the regional distribution of endothelin was determined in porcine tissue. The highest concentration of immunoreactive endothelin was observed in inner medulla of kidney (6.2 +/- 1.1 pg/mg wet weight), while the concentration in kidney cortex was very low. Immunoreactive endothelin was also found in lung in relatively high concentration. The immunoreactive endothelin in porcine lung and inner medulla of kidney was further characterized by reverse phase high performance liquid chromatography combined with radioimmunoassay.     

Cloning and sequence analysis of cDNAs for human high molecular weight and low molecular weight prekininogens. Primary structures of two human prekininogens

cDNA sequences for both human high molecular weight (HMW) and low molecular weight (LMW) prekininogens have been isolated by molecular cloning and determined by sequence analysis. The sequence determination together with the S1 nuclease mapping and RNA blot-hybridization analyses indicate that human HMW and LMW prekininogen mRNAs share an identical sequence throughout the 5'-untranslated region and the protein-coding region up to the sequence encoding the 12 amino acids distal to the bradykinin sequence, and the two mRNAs then completely diverge from each other. The signal peptide, the heavy chain (H chain), and the bradykinin moiety, which are common between the two prekininogens, consist of 18, 362, and 9 amino acids, respectively, while the light chains (L chains) of the HMW and LMW prekininogens are composed of 255 and 38 amino acids, respectively. All 17 cysteine residues present in the human and bovine H chains are located at exactly equivalent positions, indicating that the human H chain, like the bovine counterpart, can form 8 loop structures, each connected by two adjacent cysteine residues. The L chains of human and bovine kininogens differ in the protein lengths as well as in some amino acids crucial for the processing of the kininogens by kallikrein. Based upon this finding, we have discussed the molecular basis for the different modes of processing of human and bovine HMW kininogens and for the different kinetics of contact activation reactions exhibited by the two HMW kininogens.     

Evaluation of four phosphopeptide enrichment strategies for mass spectrometry-based proteomic analysis

Information about phosphorylation status can be used to prioritize and characterize biological processes in the cell. Various analytical strategies have been proposed to address the complexity of phosphorylation status and comprehensively identify phosphopeptides. In this study, we evaluated four strategies for phosphopeptide enrichment, using titanium dioxide (TiO₂) and Phos-tag ligand particles from in-gel or in-solution digests prior to mass spectrometry-based analysis. Using TiO₂ and Phos-tag magnetic beads, it was possible to enrich phosphopeptides from in-gel digests of phosphorylated ovalbumin separated by Phos-tag SDS-PAGE or in-solution serum digests, while minimizing non-specific adsorption. The tip-column strategy with TiO₂ particles enabled enrichment of phosphopeptides from in-solution digests of whole-cell lysates with high efficiency and selectivity. However, the tip-column strategy with Phos-tag agarose beads yielded the greatest number of identified phosphopeptides. The strategies using both types of tip columns had a high degree of overlap, although there were differences in selectivity between the identified phosphopeptides. Together, our results indicate that multi-enrichment strategies using TiO₂ particles and Phos-tag agarose beads are useful for comprehensive phosphoproteomic analysis.