RO 15-1788 decreases hypnotic effects of sleep deprivation

The present study investigated the effects of 5 mg, 60 mg and 120 mg of the benzodiazepine antagonist RO 15-1788 on the ability to resist sleep and on mood of sleep deprived subjects. Repeated administration of 60 and 120 mg significantly increased subjects alertness in comparison with 5 mg and placebo. The 5 mg dose had a tendency to potentiate the hypnotic effects of sleep-deprivation. The higher levels of the drug also decreased positive mood and increased negative mood, and increased the density of sleep spindles during sleep. These results are interpreted to suggest a dose dependent effect of RO 15-1788 on arousal level.     

Enzymatic Protein Carboxyl Methylation in Rat Posterior Pituitary: Neurophysins in Rapid-Turnover Pool Determine Methyl Accepting Capacity

In vitro stimulation of intact rat posterior pituitary by either veratridine or K+ depolarization results in the concomitant release of neurophysins and in a decrease (70-80%) in their carboxyl methylation as measured either with L-[methyl-3H]methionine in the intact lobes after stimulation or in their homogenates with [methyl-3H]S-adenosyl-L-methionine and purified protein carboxyl methyltransferase. A similar reduction in neurophysin methylation (60%) was observed when the arrival of newly synthesized neurophysins at the posterior pituitary was blocked by colchicine. Experimental data indicate that the reduction in neurophysin content of the lobes after 12 h of colchicine treatment (less than 7%) or after in vitro stimulation (about 10%) cannot account for the marked reduction in neurophysin methylation. The results suggest that the granule pool characterized by rapid turnover of neurophysins probably represents the major source of methyl acceptor proteins in the lobe. In spite of the marked reduction in neurophysin methyl accepting capacity observed after stimulation, there was no parallel increase in methyl accepting capacity of the released neurophysins. We propose that a neurophysin subfraction that might be associated with the membrane of releasable granules participates in the methylation reaction in situ.     

Two 19th-century chronobiologists: Thomas Laycock and Edward Smith.

This article describes the works of two 19th-century chronobiologists. Thomas Laycock (1812-1876), who held the Chair of Medicine in Edinburgh from 1855-1876, published a series of seven articles in Lancet, all dedicated to periodicities in "vital phenomena." Laycock considered the understanding of periodicities essential for the advancement of the treatment of diseases. Edward Smith (1818-1874) was a pioneer in experimental chronobiology. In his 1861 book entitled: Health and disease as influenced by daily, seasonal and other cyclical changes in the human, Smith summarized a large number of experiments in which he investigated the occurrence of periodicities in pulse rate, urine flow, urea excretion, and respiration. From his experimental results and those of others, Smith drew practical conclusions regarding patients' care, the timing of drug administration, and the design of night work.     

A Hypernychthemeral Sleep-Wake Syndrome: A Treatment Attempt

The wake and sleep-onset times of a patient with a sleep-wake cycle longer than 24 hr were recorded by the patient for 4 years. During this time, the patient found himself unable to maintain a 24-hr sleep-wake schedule. When treated with 1-2 mg clonazepam, taken nightly, he was able to become entrained to a 24-hr day. Despite entrainment of his sleep-wake cycle, the patient reported depression, lack of motivation and fatigue and chose not to continue taking the drug.     

Circular dichroism and hydrodynamic measurements of ternary protein--two ligand systems: serum albumin-bilirubin-drug or alcohol.

The effects of various ligands on bilirubin-serum albumin complexes in aqueous solution were investigated at pH 7.4 and 27 °C by circular dichroism (CD) measurements. The ligands included various penicillins, benzoic acid derivatives, and various lower aliphatic alcohols, using a molar excess of charcoal-treated human or bovine serum albumin with respect to bilirubin. In all cases investigated, significant changes in the visible-range CD spectra of the bilirubin-serum albumin complexes occurred within a certain range of added ligand concentrations. For several such ligand systems, analogous CD effects could be measured on both diluted and undiluted human blood serum or plasma. For part of the isolated albumin-ligand systems, significant dissociation of the bilirubin from the albumin was demonstrated by electrophoretic and analytical ultracentrifugation measurements, while other systems did not reveal measurable dissociation under the conditions used, indicating the formation of a ternary complex. A scheme of equilibria among all complex components is proposed, which includes both dissociation of the bilirubin and ternary complex formation in which the bilirubin conformation appears to be modified. At least two different sets of binding sites (competitive and noncompetitive) for added ligands are assumed. Values of apparent parameters describing the formation of ternary complexes from the bilirubin-albumin complex are estimated for a number of systems. Some relationships between the chemical structure of a ligand and its effect on the bilirubin-serum albumin complex are deduced. The relevance of the results obtained for the isolated protein-ligand complexes with respect to in vivo conditions is evaluated.     

Herbicide response polymorphisms in wild emmer wheat: ecological and isozyme correlations

Summary We demonstrate that the scores and frequencies of chlortoluron (CT) and metoxuron (MX) resistance and susceptible phenotypes of wild emmer wheat, Triticum dicoccoides, are correlated with ecological factors and allozyme markers. Some isozyme markers located on chromosome 6B (e.g. Adh,Est-4 and Got), which also harbours the CT and MX resistance gene, provide good genetic markers for herbicide resistance breeding. Significant correlations between herbicide and photosynthetic characters suggest that the evolution of herbicide resistance polymorphisms may be related to the process of photosynthesis in nature and predated domestication of cultivated wheat.     

Crystal structure of a type II dehydroquinate dehydratase-like protein from Bifidobacterium longum

Dehydroquinate dehydratase (DHQD) catalyzes the third step in the biosynthetic shikimate pathway. Here we identify a Bifidobacterium longum protein with high sequence homology to type II DHQDs but no detectable DHQD activity under standard assay conditions. A crystal structure reveals that the B. longum protein adopts a DHQD-like tertiary structure but a distinct quaternary state. Apparently forming a dimer, the B. longum protein lacks the active site aspartic acid contributed from a neighboring protomer in the type II DHQD dodecamer. Relating to the absence of protein–protein interactions established in the type II DHQD dodecameric assembly, substantial conformational changes distinguish the would-be active site of the B. longum protein. As B. longum possess no other genes with homology to known DHQDs, these findings imply a unique DHQD activity within B. longum.     

Elevated Plasma Homocysteine in Older Shift‐Workers: A Potential Risk Factor for Cardiovascular Morbidity

There is evidence supporting an association between shift work and cardiovascular morbidity, but the underlying mechanisms are unknown. The present paper investigated the levels of cardiovascular biochemical risk factors in shift‐workers both with (n=26) and without (n=103) sleep complaints, and in day‐workers (n=173) working in the same plant. Blood samples were taken in the morning after an overnight fast and analyzed for homocysteine, C‐reactive protein, and lipid profile. Biochemical data were compared among groups after stratifying workers by age (i.e., <40 and ≥40 yrs). Shift‐workers who complained about sleep disturbances and who were ≥40 years of age had significantly higher levels of homocysteine than did their younger counterparts—shift‐workers who did not complain of sleep disturbances and day‐workers. There were no other between‐group differences in any of the biochemical variables. The results of this investigation demonstrate an association between sleep disturbances in older shift‐workers and mild hyperhomocysteinemia. The elevated homocysteine levels may play a role in the increased rates of cardiovascular morbidity in shift‐workers, and they may have practical implications regarding the nutrition of shift‐workers.     

Modifying human thymidylate kinase to potentiate azidothymidine activation

Based on the knowledge of the crystal structures of yeast and Escherichia coli thymidylate kinases (TmpKs) and the observation that TmpK from E. coli can phosphorylate azidothymidine monophosphate (AZT-MP) much more efficiently than either the yeast or the highly homologous human enzyme, we have engineered yeast and human TmpKs to obtain enzymes that have dramatically improved AZT-MP phosphorylation properties. These modified enzymes have properties that make them attractive candidates for gene therapeutic approaches to potentiating the action of AZT as an inhibitor of human immunodeficiency virus (HIV) replication. In particular, insertion of the lid domain of the bacterial TmpK into the human enzyme results in a pronounced change of the acceptance of AZT-MP such that it is now phosphorylated even faster than TMP.     

Structural characterization of the closed conformation of mouse guanylate kinase

Guanylate kinase (GMPK) is a nucleoside monophosphate kinase that catalyzes the reversible phosphoryl transfer from ATP to GMP to yield ADP and GDP. In addition to phosphorylating GMP, antiviral prodrugs such as acyclovir, ganciclovir, and carbovir and anticancer prodrugs such as the thiopurines are dependent on GMPK for their activation. Hence, structural information on mammalian GMPK could play a role in the design of improved antiviral and antineoplastic agents. Here we present the structure of the mouse enzyme in an abortive complex with the nucleotides ADP and GMP, refined at 2.1 A resolution with a final crystallographic R factor of 0.19 (R(free) = 0.23). Guanylate kinase is a member of the nucleoside monophosphate (NMP) kinase family, a family of enzymes that despite having a low primary structure identity share a similar fold, which consists of three structurally distinct regions termed the CORE, LID, and NMP-binding regions. Previous studies on the yeast enzyme have shown that these parts move as rigid bodies upon substrate binding. It has been proposed that consecutive binding of substrates leads to "closing" of the active site bringing the NMP-binding and LID regions closer to each other and to the CORE region. Our structure, which is the first of any guanylate kinase with both substrates bound, supports this hypothesis. It also reveals the binding site of ATP and implicates arginines 44, 137, and 148 (in addition to the invariant P-loop lysine) as candidates for catalyzing the chemical step of the phosphoryl transfer.