5''-Nucleotidases in rat heart. Evidence for the occurrence of two soluble enzymes with different substrate specificities.

Chromatography of soluble proteins from rat heart on phosphocellulose columns separates two 5'-nucleotidases. The first to emerge from the column shows a preference for AMP over IMP as substrate, whereas the second shows a preference for IMP over AMP. The properties of the IMP-preferring enzyme, including the conditions under which it is eluted from phosphocellulose columns, show it to be the enzyme studied by Itoh, Oka & Ozasa [Biochem. J. (1986) 235, 847-851]. The kinetic properties of the AMP-preferring enzyme indicate that it is likely to be the enzyme responsible for the production of adenosine under conditions of hypoxia and increased work load, and with metabolic stresses such as a high load of acetate.     

Monoamine oxidase inhibitory properties of milacemide in rats

Milacemide is a glycine prodrug with reported antiepileptic antimyoclonic properties. In this study, milacemide increased "wet dog shakes" in rats pretreated with 5-Hydroxytryptophan (5-HTP) and carbidopa. Moreover, it worsened the serotonin behavior syndrome precipitated by 5-HTP and the monoamine oxidase inhibitor tranylcypromine. The serotonin syndrome was also elicited by the combination of milacemide and 5-HTP without tranylcypromine. In vitro, milacemide inhibited both monoamine oxidase A and B from the frontal cortex of rats, to a greater extent for MAO B. This drug is currently under investigation in humans as an antiepileptic agent and precautions for the consequences of monoamine oxidase inhibition should be considered when the drug is used in high doses.     

Sex determination and differentiation in organisms other than higher plants

The understanding of sex determination in general, but in particular in mammals, has been a subject of scientific speculation for a long time. It has been shown that in many vertebrate and invertebrate species, the sex of an individual is determined by the individual's chromosomal constitution. Initial studies of classical genetic searching for sex-transforming mutations and the scrupulous analyses of modified phenotypes have shed light on the mechanism(s) of sex-determination. They paved the road to successful studies at molecular level. After a brief review on sex determination in chosen model species, the “Drosophila system” is presented to exemplify a possible general principle for sex determinism.     

PI 3-kinase is a dual specificity enzyme: autoregulation by an intrinsic protein-serine kinase activity.

Phosphatidylinositol 3-kinase (PI 3-kinase) has a regulatory 85 kDa adaptor subunit whose SH2 domains bind phosphotyrosine in specific recognition motifs, and a catalytic 110 kDa subunit. Mutagenesis of the p110 subunit, within a sequence motif common to both protein and lipid kinases, demonstrates a novel intrinsic protein kinase activity which phosphorylates the p85 subunit on serine at a stoichiometry of approximately 1 mol of phosphate per mol of p85. This protein-serine kinase activity is detectable only upon high affinity binding of the p110 subunit with its unique substrate, the p85 subunit. Tryptic phosphopeptide mapping revealed that the same major peptide was phosphorylated in p85 alpha both in vivo in cultured cells and in the purified recombinant enzyme. N-terminal sequence and mass analyses were used to identify Ser608 as the major phosphorylation site on p85 alpha. Phosphorylation of the p85 subunit at this serine causes an 80% decrease in PI 3-kinase activity, which can subsequently be reversed upon treatment with protein phosphatase 2A. These results have implications for the role of inter-subunit serine phosphorylation in the regulation of the PI 3-kinase in vivo.     

New Isopropyl Chromone and Flavanone Glucoside Compounds from the Leaves of Syzygium cerasiforme (Blume) Merr. & L.M.Perry and Their Inhibition of Nitric Oxide Production

A new isopropyl chromone ( 1 ) and a new flavanone glucoside ( 2 ) together with eleven known compounds ( 3–13 ) were isolated from the leaves of Syzygium cerasiforme (Blume) Merr. & L.M.Perry. Their structures were elucidated as 5,7-dihydroxy-2-isopropyl-6,8-dimethyl-4H-chromen-4-one ( 1 ), 5,7-dihydroxyflavanone 7-O-β-D-(6′′-O-galloylglucopyranoside) ( 2 ), strobopinin ( 3 ), demethoxymatteucinol ( 4 ), pinocembrin-7-O-β-D-glucopyranoside ( 5 ), (2S)-hydroxynaringenin-7-O-β-D-glucopyranoside ( 6 ), afzelin ( 7 ), quercetin ( 8 ), kaplanin ( 9 ), endoperoxide G3 ( 10 ), grasshopper ( 11 ), vomifoliol ( 12 ), litseagermacrane ( 13 ) by the analysis of HR-ESI-MS, NMR, and CD spectral data. Compounds 1 , 2 , 5 , 6 and 10 inhibited NO production on LPS-activated RAW264.7 cells with IC₅₀ values of 12.28±1.15, 8.52±1.62, 7.68±0.87, 9.67±0.57, and 6.69±0.34 μM, respectively, while the IC₅₀ values of the other compounds ranging from 33.38±0.78 to 86.51±2.98 μM, compared to that of the positive control, N^G-monomethyl-L-arginine acetate (L-NMMA) with an IC₅₀ value of 32.50±1.00 μM.     

Novel (E)-3-(3-Oxo-4-substituted-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-N-hydroxypropenamides as Histone Deacetylase Inhibitors: Design,Synthesis and Bioevaluation

Herein, we report the design, synthesis and evaluation of novel (E)-3-(3-oxo-4-substituted-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-N-hydroxypropenamides ( 4 a – i , 7 a – g ) targeting histone deacetylases. Three human cancer cell lines were used to test the cytotoxicity of the synthesized compounds (SW620, colon; PC-3, prostate; NCI−H23, lung cancer); inhibitory activity towards HDAC; anticancer activity; as well as their impact on the cell cycle and apoptosis. As a result, compounds 4 a – i bearing the alkyl substituents seemed to be less potent than the benzyl-containing compounds 7 a – g in all biological assays. Compounds 7 e – f were found to be the most active HDAC inhibitors with IC₅₀ of 1.498±0.020 μM and 1.794±0.159 μM, respectively. In terms of cytotoxicity and anticancer assay, 7 e and 7 f also showed good activity with IC₅₀ values in the micromolar range. In addition, the cell cycle and apoptosis of SW620 were affected by compound 7 f in almost a similar manner to that of reference compound SAHA. Docking assays were carried out for analysis the binding mode and selectivity of this compound toward 8 HDAC isoforms. Overall, our data confirmed that the inhibition of HDAC plays a pivotal role in their anticancer activity.     

Chemico-structure of the organophosphatic shells of siphonotretide brachiopods

The organophosphatic shell of siphonotretide brachiopods is stratiform with orthodoxly secreted primary and secondary layers. The dominant apatitic constituents of the secondary layer are prismatic laths and rods arranged in monolayers (occasionally in cross-bladed successions), normally recrystallized as platy laminae. Sporadically distributed, interlaminar, lenticular chambers, containing apatitic meshes of laths and aggregates of plates and spherulites, probably represent degraded, localized exudations of glycosaminoglycans (GAGs) with dispersed apatite.
The shells of Helmersenia and Gorchakovia are perforated by canals with external depressions (antechambers) that possibly contained chitinous tubercles in vivo . The immature shell of Siphonotreta and most other siphonotretids is similarly perforated and pitted; but the mature part bears recumbent, rheomorphic, hollow spines that grew forward out of pits. Internally, spines pierce the shell as independent structures to terminate as pillars in GAGs chambers. Spines and pillars were probably secreted by collectives of specialized cells (acanthoblasts) within the mantle.
The shell of the oldest siphonotretide, Schizambon , is imperforate but the ventral valve has a pedicle foramen that lies forward of the posterior margin of the juvenile valve. This relationship characterizes all siphonotretides, suggesting that the pedicle, in vivo , originated within the ventral outer epithelium and not from the posterior body wall as in lingulides.     

On peptide bond formation, translocation, nascent protein progression and the regulatory properties of ribosomes. Derived on 20 October 2002 at the 28th FEBS Meeting in Istanbul.

High-resolution crystal structures of large ribosomal subunits from Deinococcus radiodurans complexed with tRNA-mimics indicate that precise substrate positioning, mandatory for efficient protein biosynthesis with no further conformational rearrangements, is governed by remote interactions of the tRNA helical features. Based on the peptidyl transferase center (PTC) architecture, on the placement of tRNA mimics, and on the existence of a two-fold related region consisting of about 180 nucleotides of the 23S RNA, we proposed a unified mechanism integrating peptide bond formation, A-to-P site translocation, and the entrance of the nascent protein into its exit tunnel. This mechanism implies sovereign, albeit correlated, motions of the tRNA termini and includes a spiral rotation of the A-site tRNA-3' end around a local two-fold rotation axis, identified within the PTC. PTC features, ensuring the precise orientation required for the A-site nucleophilic attack on the P-site carbonyl-carbon, guide these motions. Solvent mediated hydrogen transfer appears to facilitate peptide bond formation in conjunction with the spiral rotation. The detection of similar two-fold symmetry-related regions in all known structures of the large ribosomal subunit, indicate the universality of this mechanism, and emphasizes the significance of the ribosomal template for the precise alignment of the substrates as well as for accurate and efficient translocation. The symmetry-related region may also be involved in regulatory tasks, such as signal transmission between the ribosomal features facilitating the entrance and the release of the tRNA molecules. The protein exit tunnel is an additional feature that has a role in cellular regulation. We showed by crystallographic methods that this tunnel is capable of undergoing conformational oscillations and correlated the tunnel mobility with sequence discrimination, gating and intracellular regulation.