Unifying Time to Contact Estimation and Collision Avoidance across Species

The [Formula: see text]-function and the [Formula: see text]-function are phenomenological models that are widely used in the context of timing interceptive actions and collision avoidance, respectively. Both models were previously considered to be unrelated to each other: [Formula: see text] is a decreasing function that provides an estimation of time-to-contact (ttc) in the early phase of an object approach; in contrast, [Formula: see text] has a maximum before ttc. Furthermore, it is not clear how both functions could be implemented at the neuronal level in a biophysically plausible fashion. Here we propose a new framework - the corrected modified Tau function - capable of predicting both [Formula: see text]-type ("[Formula: see text]") and [Formula: see text]-type ("[Formula: see text]") responses. The outstanding property of our new framework is its resilience to noise. We show that [Formula: see text] can be derived from a firing rate equation, and, as [Formula: see text], serves to describe the response curves of collision sensitive neurons. Furthermore, we show that [Formula: see text] predicts the psychophysical performance of subjects determining ttc. Our new framework is thus validated successfully against published and novel experimental data. Within the framework, links between [Formula: see text]-type and [Formula: see text]-type neurons are established. Therefore, it could possibly serve as a model for explaining the co-occurrence of such neurons in the brain.     

Influence of 1-aminocyclohexane-1-carboxylic acid in position 2 or 3 of AVP and its analogues on their pharmacological properties.

In this study we describe the synthesis and some pharmacological properties of seven new analogues of arginine vasopressin (AVP) substituted in position 2 or 3 with 1-aminocyclohexane-1-carboxylic acid (Acc). All peptides were tested for the pressor, antidiuretic and uterotonic in vitro activities. The Acc3 modifications of AVP, dAVP, [d-Arg8]VP and [Cpa1]AVP have been found to be deleterious for interaction with all three neurohypophyseal hormone receptors, as judged from the several orders of magnitude decreased biological activities, whereas Acc2 substitution selectively altered the interaction with the receptors. Two of the new analogues, [Acc2]AVP and [Acc2, d-Arg8]AVP, are potent antidiuretic agonists. [Acc2]AVP exhibits moderate pressor agonistic activity and weak antiuterotonic properties. [Acc2, d-Arg8]AVP has been found to be a weak antagonist in the pressor and uterotonic tests. Another analogue - [Cpa1, Acc3]AVP - turned out to be a highly selective V2 agonist. This is an unexpected effect, as its parent peptide, [Cpa1]AVP is a very potent V1a receptor antagonist. This is the first Cpa1 modification to have resulted in V2 agonism enhancement. Besides providing useful information about structure-activity relationships, our results could open up new possibilities in the design of highly potent and selective V2 agonists.     

Continuous molecular evolution of protein-domain structures by single amino acid changes

Protein structures cluster into families of folds that can result from extremely different amino acid sequences [1]. Because the enormous amount of genetic information generates a limited number of protein folds [2], a particular domain structure often assumes numerous functions. How new protein structures and new functions evolve under these limitations remains elusive. Molecular evolution may be driven by the ability of biomacromolecules to adopt multiple conformations as a bridge between different folds [3-6]. This could allow proteins to explore new structures and new tasks while part of the structural ensemble retains the initial conformation and function as a safeguard [7]. Here we show that a global structural switch can arise from single amino acid changes in cysteine-rich domains (CRD) of cnidarian nematocyst proteins. The ability of these CRDs to form two structures with different disulfide patterns from an identical cysteine pattern is distinctive [8]. By applying a structure-based mutagenesis approach, we demonstrate that a cysteine-rich domain can interconvert between two natively occurring domain structures via a bridge state containing both structures. Comparing cnidarian CRD sequences leads us to believe that the mutations we introduced to stabilize each structure reflect the birth of new protein folds in evolution.     

Modulation of cell proliferation in rat liver cell cultures by new calix[4]arenes

Cell cycle progression is dependent on intracellular iron level and chelators lead to iron depletion and decrease cell proliferation. This antiproliferative effect can be inhibited by exogenous iron. In this work, we present the synthesis of new synthetic calix[4]arene podands bearing two aspartic/glutamic acid, ornithine groups or hydrazide function at the lower rim, designed as potential iron chelators. The synthesis only afforded calix[4]arenes in the cone conformation. We report their effect on cell proliferation, in comparison with the new oral chelator ICL670A (4-[3,5-bis-(2-hydroxyphenyl)-1,2,4-triazol-1-yl]-benzoic acid). The antiproliferative effect of these new compounds was studied in the rat hepatoma cell line Fao by measuring mitochondrial succinate dehydrogenase activity. Their cytotoxicity was evaluated by extracellular LDH activity. Preliminary results indicated that among all tested compounds, monohydrazidocalix[4]arene 2 which is not cytotoxic in Fao cells exhibits interesting antiproliferative activity. This effect, independent on iron depletion, remains to be further explored. Moreover, it also shows that new substituted calix[4]arenes could open the way to new valuable medicinal chemistry scaffolding.     

Heterocyclic ortho-quinones, a novel type of Photosystem II inhibitors

Members of the new chemical class of 7-substituted 6-bromo-benzo[4,5]imidazo[1,2alpha]pyridin-8,9-diones were found to be excellent inhibitors at the Q(B) site of the photosystem II D1 reaction center protein. The best inhibitors with pI(50)-values of >7 are: dimethyl-propyl, 7.05; i-pentyl, 7.36; t. butyl, 7.47; and i-propyl, 7.51. Displacement experiments with [14C]atrazine revealed that the 8,9-diones behave non-competitively in respect of Photosystem II herbicides and, hence, have to be considered as a new type of Photosystem II inhibitors. This notion is further corroborated by their inhibitory activity in D1 mutants of Chlamydomonas reinhardtii.     

A new isotopic assay for purine nucleoside phosphorylase

We have developed a new assay for purine nucleoside phosphorylase which is based on the release of tritium when [2-3H]inosine is used as the substrate and the reaction is coupled with xanthine oxidase. After the reaction is terminated, residual [2-3H]inosine is adsorbed on charcoal and the supernatant solution is assayed for radioactivity by liquid scintillation spectrometry. The new method gave results indistinguishable from those obtained by spectrophotometric determination of uric acid produced by the phosphorylase-xanthine oxidase-coupled reaction or by radioassay of chromatographically isolated [8-14C]hypoxanthine when [8-14C]inosine was used as substrate. The new method is faster than those involving chromatographic isolation of products. In comparison with spectrophotometric methods, it not only requires less manual time, but it also has the advantage that it can be used to study inhibitors whose ultraviolet absorption might interfere with spectrophotometric determination of uric acid.     

A New Non-Mendelian Genetic Element of Yeast That Increases Cytopathology Produced by M1 Double-Stranded RNA in ski Strains

The Saccharomyces cerevisiae SKI (superkiller) genes are repressors of replication of M, L-A, and L-BC double-stranded (ds) RNAs; ski strains have an increased M dsRNA copy number and, as a result, are cold-sensitive for growth at 8 degrees. Growth is normal, however, at higher temperatures. We have found a new cytoplasmic genetic element [D] (for disease) that makes M1 dsRNA-containing superkiller strains grow slowly at 30 degrees, not at all at 37 degrees, and only very poorly at 20 degrees. These growth defects require three factors: a chromosomal ski mutation, the presence of M1 dsRNA, and the presence of the new cytoplasmic factor, [D]. We have isolated mutants unable to maintain [D] (mad), at least one of which is due to mutation of a single chromosomal locus. Further, [D] can be cured by growth at 37-39 degrees. We present evidence that [D] is not M, L-A, L-BC or W dsRNAs or mitochondrial DNA, 2 mu DNA, or [psi], but [D] depends on L-A for its maintenance. We also show that [D] is distinct from [B], a cytoplasmic element that allows M1 dsRNA to be stably replicated and maintained in spite of defects in certain chromosomal MAK genes that would otherwise be necessary. [D] activity is blocked by the presence of another extrachromosomal element, called [DIN] (for [D] interference). [D] and [DIN] may be different natural variants of the same molecule.     

Effects of choline augmentation on acetylcholine synthesis and release in rat atrial minces

The uptake and acetylation of [3H]-choline, as well as the calcium-dependent release of a newly synthesized [3H]-ACh, was studied in a new rat atrial mince preparation. The hemicholinium-3-sensitive uptake and acetylation of [3H]-choline increased as [3H]-choline concentrations were elevated to 100 microM in atrial minces. In contrast, hemicholinium-3-sensitive [3H]-choline uptake was saturated with 15 microM [3H]-choline in brain synaptosomes. The increased atrial [3H]-ACh synthesized in the presence of [3H]-choline augmentation was releasable by 50 mM K+-depolarization in a 1 mM cobalt-sensitive manner. These results suggest that atrial parasympathetic activity may be more sensitive to circulating choline concentrations than brain cholinergic neurons are.     

Design,synthesis and structure-activity relationship study of piperazinone-containing thieno[3,2-d]pyrimidine derivatives as new PI3Kδ inhibitors

Two classes of piperazinone-containing thieno[3,2-d]pyrimidines were designed and synthesized as new PI3Kδ inhibitors in this study. Detailed SAR study with respect to the piperazinone substituents at the 6-position of thieno[3,2-d]pyrimidine core demonstrated that piperazinone-containing thieno[3,2-d]pyrimidines would be more potent and selective for PI3Kδ than their piperazine counterparts, which led to the discovery of several potent PI3Kδ inhibitors with comparable or better antiproliferative activity against a panel of non-Hodgkin lymphoma (NHL) cell lines as compared with idelalisib. Our study will promote the development of new PI3Kδ inhibitors based on piperazinone-containing thieno[3,2-d]pyrimidine scaffold.     

Soybean lipoxygenase-1 is not a quinoprotein

Soybean lipoxygenase-1 was reinvestigated with respect to its quinoprotein nature. It has been reported previously that soybean lipoxygenase-1 contains pyrroloquinoline quinone as the organic cofactor [1]. Because spectroscopie data were found to be inconsistent [2] with the evidence presented in [1], we sought to reproduce the published data by carefully following the procedures described in [1] and supplementing them with new analytical results. The combined data lead us to conclude that soybean lipoxygenase-1 is not a quinoprotein.     

Antiproliferative effect on HepaRG cell cultures of new calix[4]arenes

Cell cycle progression is dependent on the intracellular iron level, and chelators lead to iron depletion and decrease cell proliferation. This antiproliferative effect can be inhibited by exogenous iron. In this work, we present the synthesis of new synthetic calix[4]arene podands bearing alkyl acid and alkyl ester groups at the lower rim, designed as potential iron chelators. We report their effect on cell proliferation, in comparison with the new oral chelator ICL670 (4-[3,5-bis-(2-hydroxyphenyl)-1,2,4-triazol-1-yl]-benzoic acid). The antiproliferative effect of these new compounds was studied in human hepatocarcinoma HepaRG cell cultures using the MTT assay. Their cytotoxicity was evaluated by extracellular LDH activity. Preliminary results indicate that their antiproliferative effect is due to their cytotoxicity. The efficiency of these compounds, comparable to that of ICL670, was independent of iron depletion. This effect remains to be further explored. Moreover, it also shows that novel substituted calix[4]arenes could open the way to new valuable medicinal chemistry scaffolding.     

Specific tritium labeling of glucosyl- and galactosylceramides at the 6-position of the carbohydrate moiety using CrO3-graphite

A new procedure for introducing tritium into the carbohydrate portions of glucosyl- and galactosylceramides was developed using a new catalyst, CrO3-graphite, which specifically oxidizes the primary alcohol group to the aldehyde. About 10% of the glycolipid was converted to the aldehyde and the aldehyde produced was then reduced back to the original form with KB3H4. After methanolysis, more than 96.7% of the radioactivities of [3H]glucosyl- and [3H]galactosylceramides were found to be located in the carbohydrate portions, and the specific activities of the [3H]galactosyl- and [3H]glucosylceramides were 2.08 to 4.30 X 10(4) cpm/nmol, which could be increased greatly by purifying the aldehydes and reducing them with KB3H4. In addition, beta-galactosidase activity was successfully determined with [3H]galactosylceramide as the enzyme substrate; the Km was 18.73 mM and the Vmax was 11.63 nmol/mg/h, indicating that no significant structural modification occurs during the oxidation.     

Isolation and identification of eight microcystins from thirteen Oscillatoria agardhii strains and structure of a new microcystin.

Microcystins (cyclic heptapeptide hepatotoxins), isolated from 13 freshwater Oscillatoria agardhii strains from eight different Finnish lakes by high-performance liquid chromatography, were characterized by amino acid analysis, fast atom bombardment mass spectrometry (FABMS), and tandem FABMS (FABMS/collisionary-induced dissociation/MS). All strains produced two to five different microcystins. In total, eight different compounds, of which five were known microcystins, were isolated. The known compounds identified were [D-Asp3]MCYST (microcystin)-LR, [Dha7]MCYST-LR, [D-Asp3]MCYST-RR, [Dha7]MCYST-RR, and [D-Asp3,Dha7]MCYST-RR. This is the first time that isolation of these toxins from Oscillatoria spp., with the exception of [D-Asp3]MCYST-RR, has been reported. Three of the strains produced a new microcystin, and the structure was assigned as [D-Asp3,Mser7]MCYST-RR. The structures of two new microcystins, produced as minor components by one Oscillatoria strain, could not be determined because of the small amounts isolated from the cells. Four strains produced [Dha7]MCYST-RR as the main toxin, but [D-Asp3]MCYST-RR was clearly the most abundant and most frequently occurring toxin among these isolates of O. agardhii.     

Genetic selection of short peptides that support protein oligomerization in vivo

An important goal in protein engineering is to control associations between designed proteins. This is most often done by fusing known, naturally occurring oligomerization modules, such as leucine zippers [1] [2] [3], to the proteins of interest [4] [5] [6]. It is of considerable interest to design or discover new oligomerization domains that have novel binding specificities [7] [8] [9] [10] [11] in order to expand the 'toolbox' of the protein engineer and also to eliminate associations of the designed proteins with endogenous factors. We report here a simple genetic selection scheme through which to search libraries for peptides that are able to mediate homodimerization or higher-order self-oligomerization of a protein in vivo. We found several peptides that support oligomerization of the lambda repressor DNA-binding domain in Escherichia coli cells, some of them as efficiently as the endogenous dimerization domain or the GCN4 leucine zipper. Many are very small, comprising as few as six residues. This study strongly supports the notion that peptide sequence space is rich in small peptides, which might be useful in protein engineering and other applications.     

Pyrrolo- and pyrazolo-[3,4-e][1,2,4]triazolo[1,5-c]pyrimidines as adenosine receptor antagonists

The discovery and development of adenosine receptor antagonists have represented for years an attractive field of research from the perspective of identifying new drugs for the treatment of widespread disorders such as inflammation, asthma and Parkinson's disease. The present work can be considered as an extension of our structure-activity relationship studies on the pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidine (PTP) nucleus, extensively investigated by us as a useful template, in particular, for the identification of A(2A) and A(3) adenosine receptor antagonists. In order to explore the role of the nitrogen at the 7-position, we performed a new synthetic strategy for the preparation of pyrrolo[3,4-e][1,2,4]triazolo[1,5-c]pyrimidine derivatives which can be considered as 7-deaza analogues of the parent PTPs. We also synthesised a novel series of pyrazolo[3,4-e][1,2,4]triazolo[1,5-c]pyrimidines as junction isomers of the reference compounds. In both cases we obtained some examples of potent antagonists (K(i) in the low nanomolar range) with variable selectivity profiles in relation to the nature of substituents introduced at the C(5)-, N(8)- and/or N(9)-positions. The pyrrolo-triazolo-pyrimidine derivative 9b appeared to be a potent A(3) adenosine receptor antagonist (K(i)=10 nM) with good selectivity over hA(1) (74-fold) and hA(2A) (20-fold) adenosine receptors combined with low activity at the hA(2B) subtype (IC(50)=906 nM). Moreover, some examples of high-affinity A(1)/A(2A) dual antagonists have been identified in both series. This work constitutes a new and important contribution for the comprehension of the interaction between PTPs and adenosine receptors.     

13-Oxomyricanol,a new [7.0]-metacyclophane from Myrica nagi

A new [7.0]-metacyclophane has been isolated from the root bark of Myrica nagi. On the basis of spectroscopy the constitution of the new compound is proposed to be 13-oxomyricanol.A new [7.0]-metacyclophane has been isolated from the root bark of Myrica nagi. On the basis of spectroscopy the constitution of the new compound is proposed to be 13-oxomyricanol.     

从植物共生菌宏基因组文库筛选新的生物催化酶基因

【目的】通过建立宏基因组文库的高通量保存与基于探针洗脱的多次膜杂交筛选方法,从植物共生菌宏基因组文库筛选具有生物催化潜力的新酶基因。【方法】首先根据滴度将初始文库噬菌体包装颗粒感染到EPI300-T1R E.coli,过夜培养后对应保存于96孔板;提取粘粒进行文库的杂交筛选。【结果】描述的洗脱条件可完全去除尼龙膜上与靶DNA结合的探针,并且尼龙膜上的靶DNA至少可用于7次探针杂交,从而明显提高宏基因组文库的筛选效率。【结论】以Enoate reductase(ER)和短链脱氢酶(SDR)的同源基因片段为探针,运用该方法经两轮筛选获得候选单克隆并进行了部分粘粒的测序,发现了新的ER和SDR同源基因,并克隆到相应的全长基因序列用于后续的表达与酶化学研究。     

Evidence for a new Z-type left-handed DNA helix: properties of Z(WC)-DNA.

The structure of Z-DNA, currently accepted as a model for all left-handed DNAs, fails to provide convincing explanations for at least four well established properties of left-handed DNA polymers in solution. However, the major discrepancies between theory and experiment are resolved by the structure presently proposed for Z[WC]-DNA, a new left-handed, zig-zag double helix with Watson-Crick-type backbone directions. Structural features of Z[WC]-DNA include the presence of an additional H-bond between each guanine N2-amino group and an adjacent phosphate oxygen, the capacity to form four-stranded, base-matched complexes that should readily precipitate from solution, and backbone progressions that are the same as B-DNA (opposite to Z-DNA). However, since Z[WC]-DNA and Z-DNA have many parameters in common, they could be difficult to distinguish in a majority of existing experiments. In view of the close relationship of the new helix to B-DNA, which allows a relatively unhindered right-to-left transition in handedness, Z[WC]-DNA is theorized to be the left-handed structure preferentially generated in vivo by the torque available in naturally occurring DNA supercoils.     

Weighting improves the "new Haseman-Elston" method

Elston et al. [Genet Epidemiol, in press] apply the results of Wright [Am J Hum Genet 1997;60:740-742] and Drigalenko [Am J Hum Genet 1998;63:1242-1245] to extend the traditional Haseman-Elston regression scheme [Haseman and Elston, Behav Genet 1972;2:3-19] to include not only linkage information contained in the sib pair's squared difference, but also information in their mean-corrected squared sum. The new algorithm detects linkage to a quantitative trait locus by modelling sib pair trait covariance as a function of identity-by-descent status. We demonstrate why this new estimator is suboptimal and can in some cases be inferior to the original Haseman-Elston method. We also describe a simple approach to estimation which improves on this new Haseman-Elston method by incorporating variance-based weights into the test statistic while staying within the linear modelling framework. In support of our theoretical claim, we conduct both a sib pair simulation and an application to GAW 10 sib pair data showing that our new estimator is superior to both the old and new Haseman-Elston schemes currently implemented in the analysis package S.A.G.E. 4.0.     

Proteinase inhibitors and activators strategic targets for therapeutic intervention.

The contributions in this meeting represent the state of the art in study of proteinases and their biological and therapeutic regulation. The near future of proteinase research will be fashioned by the new horizons of proteomics research [61], adding substance to the genomic data. Even new approaches in drug discovery, such as combinatorial chemistry, impact upon the understanding of the proteinase function, as with the discovery of a novel allosteric exosite in FVIIa by probing with libraries of ligands [62]. What is without question is that proteinases will remain at the forefront of understanding and intervention in human biochemistry and human disease pathology.