Protein synthesis in rabbit reticulocytes. A study of the roles of Co-eIF-2, Co-eIF-2A80, and GDP in peptide chain initiation

The roles of Co-eIF-2, Co-eIF-2A80, and GDP in ternary complex and Met-tRNAf X 40 S initiation complex formation were studied. 1) Partially purified eukaryotic initiation factor 2 (eIF-2) (50% pure) preparations contained 0.4-0.6 pmol of bound GDP/pmol of eIF-2. eIF-2 purity was calculated from ternary complex formation in the absence of Mg2+ and in the presence of excess Co-eIF-2. 2) In the absence of Mg2+, approximately 30% of the potentially active eIF-2 molecules formed ternary complexes, and both Co-eIF-2 and Co-eIF-2A80 were equally effective in full activation of the eIF-2 molecules for ternary complex formation. 3) In the presence of Mg2+, approximately 10% of the potentially active eIF-2 molecules formed ternary complexes in the absence of ancillary factors, and the ancillary factors Co-eIF-2A80 and Co-eIF-2 raised the incorporation to 20 and 50% of the eIF-2 molecules, respectively. 4) In the absence of Mg2+, [3H]GDP in preformed eIF-2 X [3H]GDP was readily displaced by GTP during ternary complex formation. 5) In the presence of Mg2+, [3H]GDP remained tightly bound to eIF-2 and ternary complex formation was inhibited. Co-eIF-2, but not Co-eIF-2A80, was effective in promoting [3H]GDP displacement and the former was more effective in promoting ternary complex formation than the latter. 6) eIF-2 X [3H]GDP was converted to eIF-2 X [3H] GTP by incubation in the presence of nucleoside-5'-diphosphate kinase and ATP, but the eIF-2 X [3H]GTP thus formed did not bind Met-tRNAf in the presence of Mg2+ and required exogeneous addition of Co-eIF-2 and GTP for ternary complex formation and GTP displacement. 7) In the absence of Mg2+, the increased ternary complex formed in the presence of eIF-2 X [3H] GDP and Co-eIF-2A80 (with accompanying loss of [3H] GDP) was inactive in a subsequent reaction, which involves Met-tRNAf transfer to 40 S ribosomes (in the presence of Mg2+), and required trace amounts of Co-eIF-2 for such activity. Based on the above observations, we have suggested a two-step activation of eIF-2 molecules by the Co-eIF-2 protein complex for functional ternary complex formation. One of these steps involves the Co-eIF-2A component of Co-eIF-2. This activation results in stimulated Met-tRNAf binding to eIF-2 and is most apparent in the absence of Mg2+ and with aged eIF-2 molecules.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of piracetam, a nootropic agent, on experimental gastric ulcers in rat

Piracetam, used clinically for cognitive disorders, was found to have significant anti-ulcerogenic activity against immobilization stress- and aspirin-induced gastric ulcers in rats. The anti-ulcer effect of piracetam was exerted by augmentation of mucosal resistance. This was indicated by the significant attenuation of the decrease in total carbohydrate: protein ratio induced by aspirin. It also reversed the marked increase in gastric juice protein and DNA induced by aspirin, indicating that piracetam attenuated the augmented mucosal cell exfoliation induced by the ulcerogen. The drug also increased gastric mucosal serotonin concentrations. Piracetam, thus appears to have a profile of activity associated with cytoprotective agents.     

Membrane perturbation through novel cell-penetrating peptides influences intracellular accumulation of imatinib mesylate in CML cells

Chronic myeloid leukemia is a stem cell disease with the presence of Philadelphia chromosome generated through reciprocal translocation of chromosome 9 and 22. The use of first- and second-generation tyrosine kinase inhibitors has been successful to an extent. However, resistance against such drugs is an emerging problem. Apart from several drug-resistant mechanisms, drug influx/efflux ratio appears to be one of the key determinants of therapeutic outcomes. In addition, intracellular accumulation of drug critically depends on cell membrane fluidity and lipid raft dynamics. Previously, we reported two novel cell-penetrating peptides (CPPs), namely, cationic IR15 and anionic SR11 present in tryptic digest of Abrus agglutinin. Here, the potential of IR15 and SR11 to influence intracellular concentration of imatinib has been evaluated. Fluorescent correlation spectroscopy and lifetime imaging were employed to map membrane fluidity and lipid raft distribution following peptide-drug co-administration. Results show that IR15 and SR11 are the two CPPs which can modulate membrane fluidity and lipid raft distribution in K562 cells. Both IR15 and SR11 significantly reduce the viability of CML cells in the presence of imatinib by increasing the intracellular accumulation of the drug.     

A kinetic model for microbial growth on solid hydrocarbons

A kinetic model has been presented to explain the growth of microorganism on solid hydrocarbons. The model is based on the assumption that metabolite produced by the growing cells helps the dissolution of the solid substrate in the aqueous medium. The linear behavior of the growth curve predicted by the model is verified experimentally.     

Swertane triterpenoids from Swertia chirata

Two novel triterpenes belonging to swertane skeleton, besides gammacer-16-en-3β-ol and 21αH-hop- 22(29)-en-3β-ol, of rare occurrence have been isolated from Swertia chirata, along with some common triterpenoids. Their structures were established on the basis of spectral and chemical evidence.     

Discovery of pyrazole carboxylic acids as potent inhibitors of rat long chain L-2-hydroxy acid oxidase

Long chain L-2-hydroxy acid oxidase 2 (Hao2) is a peroxisomal enzyme expressed in the kidney and the liver. Hao2 was identified as a candidate gene for blood pressure (BP) quantitative trait locus (QTL) but the identity of its physiological substrate and its role in vivo remains largely unknown. To define a pharmacological role of this gene product, we report the development of selective inhibitors of Hao2. We identified pyrazole carboxylic acid hits 1 and 2 from screening of a compound library. Lead optimization of these hits led to the discovery of 15-XV and 15-XXXII as potent and selective inhibitors of rat Hao2. This report details the structure activity relationship of the pyrazole carboxylic acids as specific inhibitors of Hao2.