Isolation and characterization of 5-carbamoylmethyluridine and 5-carbamoylmethyl-2-thiouridine from human urine

Two new modified uracil nucleosides, 5-carbamoylmethyuridine (ncm5U, I) and 5-carbamoylmethyl-2-thiouridine (ncm5s2U, II) were isolated from a 24 hr collection of a normal human urine. The structures were assigned on the basis of UV, NMR and mass spectral data and confirmed by comparison of the spectral data and HPLC mobilities with those of authentic samples. On the basis of experimental data it appears possible that 5-carbamoylmethyl-2-thio-uridine (ncm5s2U, II) may be a degradation product produced from a labile precursor by the chemical treatments during the isolation procedure. However, the other nucleoside (ncm5U,I) certainly appears to be of metabolic origin and was also found in the urines of one chronic myelogenous leukemia and one lung carcinoma patient. Abbreviations used are: tRNA-transfer ribonucleic acid, TMS-trimethylsilyl, RP-HPLC--reverse phase high performance liquid chromatography, EI--electron impact, cm5U-5-carboxymethyluridine, mcm5U-5-methoxycarbonylmethyluridine, cm5s2U-5-carboxymethyl-2-thiouridine, mcm5s2U-5-methoxycarbonylmethyl-2-thiouridine, t6A-9-beta-D-ribofuranosyl-[N(purin-6-yl)carbamoyl]-1-threonine, C-cytidine, acp3u-3-(3-amino-3-carboxypropyl)uridine, AICR-aminoimidazole carboxamide riboside, alpha-4-PCNR & beta-4-PCNR-9-alpha-D-(or beta-D)-ribofuranosyl-pyridin-4-one-3-carboxamide, H x 7R-7-beta-D-ribofuranosyl hypoxanthine, m3U-3-methyluridine, m1I-1-methylinosine, m1G-1-methylguanosine, DI-5'-deoxyinosine, dms5OA-5'-deoxy-5'-methylthioadenosine sulfoxide, m2(2)G-N2-dimethylguanosine, psi-psi-uridine, A-adenosine, I-inosine, CML-chronic myelogenous leukemia mam5s2U-5-methylaminomethyl-2-thiouridine, ncm5U-5-carbamoylmethyluridine, ncm5s2U-5-carbamoylmethyl-2-thiouridine, UV-ultraviolet, NMR-nuclear magnetic resonance, HPLC-high performance liquid chromatography, GC-MS-gas chromatography-mass spectrometry.     

Histidine kinases: diversity of domain organization

Histidine kinases play a major role in signal transduction in prokaryotes for the cellular adaptation to environmental conditions and stresses. Recent progress in the three-dimensional structure determination of two representative members of histidine kinases, EnvZ (class I) and CheA (class II), has revealed common structural features, as well as a kinase catalytic motif topologically similar to those of the ATP-binding domains of a few ATPases. They have also disclosed that there are significant differences in domain organization between class I and II histidine kinases, possibly reflecting their distinct locations, functions and regulatory mechanisms. In spite of this diversity, both class I and II histidine kinases use similar four-helix bundle motifs to relay phosphoryl groups from ATP to regulatory domains of response regulators. The previously known so-called transmitter domain of histidine kinase is further dissected into two domains: a CA (Catalytic ATP-binding) domain and a DHp (Dimerization Histidine phosphotransfer) domain for class I, or a CA domain and an HPt (Histidine-containing Phosphotransfer) domain for class II histidine kinases. From a comparative analysis of the CA domains of EnvZ, CheA and their ATPase homologues, the core elements of the CA domain have been derived. The apparent resemblance between DHp and HPt domains is only superficial, and significant differences between them are discussed.     

Kinetic asymmetry as a key source of functional diversity in biochemical networks

From the analysis of the dynamic properties of various symmetric and asymmetric kinetic schemes, the present report demonstrates that all kinetic schemes, which can be hypothetically divided into two equal halves about an axis of mirror symmetry, are endowed with structural metastability under mass-closed conditions. In mass-closed symmetric schemes, absolute symmetry in reaction conditions in two halves is essential for the occurrence of ordered dynamic behaviour. Even an infinitesimal deviation from the symmetry relations instantaneously drives such systems from limit-cycles to turbulence. Reaction schemes with no axes of symmetry may exhibit a large variety of complex, structurally stable temporal order for wide ranges of values of system parameters and variables. Kinetic asymmetry, therefore, may confer to biochemical networks the functional diversity as well as stability against environmental perturbations.     

Biodegradation of n-alkylcycloalkanes and n-alkylbenzenes via new pathways in Alcanivorax sp. strain MBIC 4326

The degradation of long-chain n-alkylbenzenes and n-alkylcyclohexanes by Alcanivorax sp. strain MBIC 4326 was investigated. The alkyl side chain of these compounds was mainly processed by beta-oxidation. In the degradation of n-alkylcyclohexanes, cyclohexanecarboxylic acid was formed as an intermediate. This compound was further transformed to benzoic acid via 1-cyclohexene-1-carboxylic acid.     

The regions of securin and cyclin B proteins recognized by the ubiquitination machinery are natively unfolded

The proteins securin and cyclin B are destroyed in mitosis by the ubiquitin/proteasome system. This destruction is important to mitotic progression. The N-terminal regions of these proteins contain the sequence features recognized by the ubiquitination system. We have demonstrated using circular dichroism and 1-D and 2-D nuclear magnetic resonance that these rather substantial regions are natively unfolded. Based on these findings, we propose a model that helps to explain previously enigmatic observations.     

Sublethal temperature stress in juvenile Labeo rohita (Ham-Buch.) and Rita rita (Ham.): some physiological changes

Juveniles of fish L. rohita and R. rita subjected to a rapid (5 min) sublethal temperature increase from 28 to 35 degrees C showed significant increase in cortisol and decrease in interrenal ascorbic acid. Hypercholesterolemia, hyperglycemia and hyperlactemia were also evident accompanied by increased blood haemoglobin and haematocrit and stable protein levels. Compensatory responses were initiated within 72 hr in both the fishes. R. rita recovered more quickly indicating it to be more resistant to the heat stress than L. rohita. Hence fishes subjected to sublethal temperature stress should be given a metabolic recovery period of 72 hr prior to further stress being applied.     

Potentiality of a new compound for in vitro differentiation between halophilic and non-halophilic vibrios

Sensitivity of 21 halophilic vibrios and 16 clinical isolates of non-halophilic vibrios was determined against a new possible antivibrio agent, a pyrimidine analogue, 4, 6-dimethylpyrimidine -2-thiol (4,6-DMPT). It appeared to be a vibriocidal agent, having a mean MIC and MBC of 32 microg/ml for halophilic strains and 64 microg/ml for non-halophilic strains and an LD50 of 300 mg/Kg body weight of mice. Thus, 4,6-DMPT may help an in vitro distinction between halophilic and non-halophilic vibrios. Sensitivity of these strains was also studied with respect to pteridine, crystal violet and Tween 80 hydrolysis as further markers distinguishing between these 2 groups which could also be differentiated by their growth on TCBS or/and CLED media.