Evidence for essential catalytic determinants for human erythrocyte pyrimidine 5′-nucleotidase

Human erythrocyte pyrimidine 5′-nucleotidase, PN-I, catalyzes the dephosphorylation of pyrimidine nucleoside monophosphates. The enzyme also possesses phosphotransferase activity, transferring phosphate groups between pyrimidine nucleoside monophosphates and various pyrimidine nucleosides. Deficiency of the enzyme activity is associated with a hemolytic anemia. PN-I cDNA has been expressed in Escherichia coli, yielding a fully active recombinant enzyme, which was purified to homogeneity and extensively characterized. Multiple sequence alignment of PN-I and homologues proteins revealed the existence of conserved regions, whose importance in catalysis was examined by performing experiments designed to intercept covalent intermediates as strongly suggested by our previous kinetic studies. Furthermore, a functional analysis of the enzyme was carried out through site-directed mutagenesis designed on the basis of the sequence of the identified conserved regions as well as mutations observed in PN-I-deficient patients.Received 25 March 2005; received after revision 3 May 2005; accepted 13 May 2005     

Which strategy for a protein crystallization project?

The three-dimensional, atomic-resolution protein structures produced by X-ray crystallography over the past 50+ years have led to tremendous chemical understanding of fundamental biochemical processes. The pace of discovery in protein crystallography has increased greatly with advances in molecular biology, crystallization techniques, cryocrystallography, area detectors, synchrotrons and computing. While the methods used to produce single, well-ordered crystals have also evolved over the years in response to increased understanding and advancing technology, crystallization strategies continue to be rooted in trial-and-error approaches. This review summarizes the current approaches in protein crystallization and surveys the first results to emerge from the structural genomics efforts.Received 1 July 2003; received after revision 6 August 2003; accepted 22 August 2003     

Triadin: a multi-protein family for which purpose?

Triadin is a protein first identified as a member of the muscle calcium release complex, involved in calcium release for muscle contraction. However, its precise function in this complex is still undefined. Recently, triadin has been shown to be a multi-protein family, with different distribution of the various splice variants within the sarcoplasmic reticulum, raising the possibility of multiple functions for this family of polypeptides. Such functions may include involvement in excitation-contraction coupling, in triad targeting, in structural function or in muscle differentiation. The putative role(s) of triadin(s) will be discussed here.Received 5 May 2004; received after revision 4 June 2004; accepted 7 June 2004     

γδ T-APCs: a novel tool for immunotherapy?

The series of seminal articles in this book clearly illustrate the multi-functional nature of γδ T cells. Some of the functions correlate with the tissue tropism of distinct γδ T cell subsets whereas others appear to result from oligoclonal selection. Here, we discuss the antigen-presenting cell (APC) function of the major subset of circulating γδ T cells, Vγ9/Vδ2 T cells, present in human blood. During tissue culture, Vγ9/Vδ2 T cells uniformly respond to a class of non-peptide antigens, so-called prenyl pyrophosphates, derived from stressed host cells or from microbes. It is this feature that distinguishes human (and primate) Vγ9/Vδ2 T cells from αβ and γδ T cells of all other species and that forms the basis for detailed studies of human Vγ9/Vδ2 T cells. One of the consequences of Vγ9/Vδ2 T cell activation is the rapid acquisition of APC characteristics (γδ T-APCs) reminiscent of mature dendritic cells (DCs). In the following discussion, we will discriminate between the potential use of γδ T-APCs as a cellular vaccine in immunotherapy and their role in anti-microbial immunity. Exploiting the APC function in γδ T-APCs represents a true novelty in current immunotherapy research and may lead to effective, anti-tumor immunity in cancer patients.     

The structure of human interferon-β: implications for activity

Interferons (IFNs) are potent extracellular protein mediators of host defence and homoeostasis. This article reviews the structure of human IFN-β (HuIFN-β), in particular in relation to its activity. The recently determined crystal structure of HuIFN-β provides a framework for understanding of the mechanism of differentiation of type I IFNs by their common receptor. Insights are generated by comparison with the structures of other type I IFNs and from the interpretation of existing mutagenesis data. The details of the observed carbohydrate structure, together with biochemical data, implicate the glycosylation of HuIFN-β, which is uncommon among type I IFNs, as an important factor in the solubility, stability and, consequently, activity of the protein. Finally, these structural implications are discussed in the context of the clinical use of HuIFN-β. Received 12 June 1998; received after revision 16 July 1998; accepted 16 July 1998     

KIF5C: A new binding partner for protein kinase CK2 with a preference for the CK2α’ subunit

Protein kinase CK2 is a highly conserved serine/threonine kinase that is ubiquitously expressed in eukaryotic cells. CK2 is a constitutively active tetrameric enzyme composed of two catalytic α and/or α’-subunits and two regulatory β-subunits. There is increasing evidence that the individual subunits may have independent functions and that they are asymmetrically distributed inside the cell. To gain a better understanding of the functions of the individual subunits, we employed a yeast-two-hybrid screen with CK2α and CK2α’. We identified the motor neuron protein KIF5C as a new binding partner for CK2. The interaction found in the yeast-two-hybrid screen was confirmed by co-sedimentation analysis on a sucrose density gradient and by co-immunoprecipitation analysis. Pull-down experiments and surface plasmon resonance spectrometry revealed a direct binding of KIF5C to CK2α’. Co-localization studies with neuroblastoma cells, bone marrow and with primary neurons confirmed the biochemical analysis that KIF5C preferentially bound to CK2α’. Received 8 August 2008; received after revision 3 November 2008; accepted 4 November 2008     

Babbage’s guidelines for the design of mathematical notations

The design of good notation is a cause that was dear to Charles Babbage’s heart throughout his career. He was convinced of the “immense power of signs” (1864, 364), both to rigorously express complex ideas and to facilitate the discovery of new ones. As a young man, he promoted the Leibnizian notation for the calculus in England, and later he developed a Mechanical Notation for designing his computational engines. In addition, he reflected on the principles that underlie the design of good mathematical notations. In this paper, we discuss these reflections, which can be found somewhat scattered in Babbage’s writings, for the first time in a systematic way. Babbage’s desiderata for mathematical notations are presented as ten guidelines pertinent to notational design and its application to both individual symbols and complex expressions. To illustrate the applicability of these guidelines in non-mathematical domains, some aspects of his Mechanical Notation are also discussed.     

The landing zone – Ground for model transfer in chemistry

This essay proposes a new notion - the landing zone - in order to identify conceptual features that allow modelers to transfer mathematical tools across disciplinary borders. This discussion refers to the transferable models as ‘templates’. Templates are functions, equations, or computational methods that are capable of being generalized from a particular subject matter. There are formal and conceptual prerequisites for the transfer of a template to a new domain. A landing zone is an ontology that contributes to the satisfaction of these conditions for successful transfer. This paper presents a case study on a model in chemistry - the Quantum Theory of Atoms in Molecules (QTAIM) - that makes use of transferred templates from physics - the virial theorem and the wave function. The landing zone in this case is a new ontological notion, that of the topological atom, which prepares ground for the use of the virial theorem and the wave function in chemistry. The virial theorem requires that there exists in-principle stability to the system that it represents, and the wave function requires transformation in its representation that is justified. The ontology of QTAIM - the landing zone for these templates - grounds the scientific use of these templates in the context of chemistry.     

A linear benchmark for forecasting GDP growth and inflation?

Predicting the future evolution of GDP growth and inflation is a central concern in economics. Forecasts are typically produced either from economic theory‐based models or from simple linear time series models. While a time series model can provide a reasonable benchmark to evaluate the value added of economic theory relative to the pure explanatory power of the past behavior of the variable, recent developments in time series analysis suggest that more sophisticated time series models could provide more serious benchmarks for economic models. In this paper we evaluate whether these complicated time series models can outperform standard linear models for forecasting GDP growth and inflation. We consider a large variety of models and evaluation criteria, using a bootstrap algorithm to evaluate the statistical significance of our results. Our main conclusion is that in general linear time series models can hardly be beaten if they are carefully specified. However, we also identify some important cases where the adoption of a more complicated benchmark can alter the conclusions of economic analyses about the driving forces of GDP growth and inflation. Copyright © 2008 John Wiley & Sons, Ltd.     

Water or ice?--the challenge for invertebrate cold survival

The ecophysiology of cold tolerance in many terrestrial invertebrate animals is based on water and its activity at low temperatures, affecting cell, tissue and whole organism functions. The normal body water content of invertebrates varies from 40 to 90% of their live weight, which is influenced by water in their immediate environment, especially in species with a water vapour permeable cuticle. Water gain from, or loss to, the surrounding atmosphere may affect animal survival, but under sub-zero conditions body water status becomes more critical for overwinter survival in many species. Water content influences the supercooling capacity of many insects and other arthropods. Trehalose is known to maintain membrane integrity during desiccation stress in several taxa. Dehydration affects potential ice nucleators by reducing or masking their activity and a desiccation protection strategy has been detected in some species. When water crystallises to ice in an animal it greatly influences the physiology of nearby cells, even if the cells remain unfrozen. A proportion of body water remains unfrozen in many cold hardened invertebrates when they are frozen, which allows basal metabolism to continue at a low level and aids recovery to normal function when thawing occurs. About 22% of total body water remains unfrozen from calculations using differential scanning calorimetry (compared with ca 19% in food materials). The ratio of unfrozen to frozen water components in insects is 1:4 (1:6 for foods). Such unfrozen water may aid recovery of freezing tolerant species after a freezing exposure. Rapid changes in cold hardiness of some arthropods may be brought about by subtle shifts in body water management. It is recognised that cold tolerance strategies of many invertebrates are related to desiccation resistance, and possibly to mechanisms inherent in insect diapause, but the role of water is fundamental to them all. Detailed experimental studies are needed to provide information which will allow a more complete and coherent understanding of the behaviour of water in biological systems and aid the cryopreservation of a wide range of biological material.     

A radioimmunoassay for dopamine-β-hydroxylase using a glass-bound antibody

Résumé Les auteurs décrivent une méthode radioimmunologique de dosage de la DH utilisant un anticorps lié de façon covalente à des billes de verre. La technique a été appliquée à la mesure de la DH dans les surrénales et différentes régions du cerveau de buf. Les principaux avantages de cette méthode sont sa relative facilité d'emploi, sa précision et sa haute spécificité.

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This work was supported by USPHS Grants Nos. MH-02717, NS-06801 and CA-02071 and NSF Grant No. GB-27603.Acknowledgements: The authors wish to express their appreciation for the technical assistance of Susan Kadner and William P. Vann.     

Age-related lysosomal dysfunction: an unrecognized roadblock for cobalamin trafficking?

Vitamin-B₁₂ is a generic term for corrinoid compounds that exhibit the biological activity of cyanocobalamin and are collectively referred to as cobalamins. Methylcobalamin and 5-deoxyadenosylcobalamin are the active cobalamins in human metabolism. Cobalamin plays a crucial role in the maintenance of homocysteine and methylmalonyl-CoA homeostasis and is required for erythrocyte formation and DNA synthesis. Data from human and animal studies indicate that cobalamin deficiency impairs neuronal function; a process that is thought to contribute to age-related cognitive decline and dementia. Cobalamin deficiency also results in dysfunction of the peripheral nervous system; among other disorders. Although there is a detailed understanding of the biochemical pathways that are perturbed in cobalamin deficiency, the mechanisms underlying age-related dyshomeostasis in such pathways remain to be addressed. Because cobalamin utilization is dependent on its efficient transit through lysosomes, and mounting evidence indicates that lysosomal function deteriorates in aging long-lived post-mitotic cells such as neurons, in the present article we review published data that supports the proposition that impaired lysosomal processing of cobalamin may play a significant role in age-related (neuro) degenerative diseases.     

Unsuitability of urethane anesthetized rats for testing potential β-adrenoreceptor blockers

Summary Isoprenaline induced tachycardia in urethane, but not sodium barbital anesthetized rats depends upon resting heart rate values. This makes urethane anesthesia unsuitable for testing -blockers.     

siRNA-mediated knock-down of NOX3: therapy for hearing loss?

Cisplatin is a widely used chemotherapeutic agent that causes significant hearing loss. Previous studies have shown that cisplatin exposure is associated with increase in reactive oxygen species (ROS) in the cochlea. The inner ear expresses a unique isoform of NADPH oxidase, NOX3. This enzyme may be the primary source of ROS generation in the cochlea. The knockdown of NOX3 by pretreatment with siRNA prevented cisplatin ototoxicity, as demonstrated by preservation of hearing thresholds and inner ear sensory cells. Trans-tympanic NOX3 siRNA reduced the expression of NOX3 and biomarkers of cochlear damage, including transient receptor vanilloid 1 (TRPV1) channel and kidney injury molecule-1 (KIM-1) in cochlear tissues. In addition, siRNA against NOX3 reduced apoptosis as demonstrated by TUNEL staining, and prevented the increased expression of Bax and abrogated the decrease in Bcl2 expression following cisplatin administration. Trans-tympanic administration of siRNA directed against NOX3 may provide a useful method of attenuating cisplatin ototoxicity. In this paper, we review recent publications dealing with the role of NOX3 in ototoxicity and the effects of siRNA against cisplatin-induced hearing loss.     

Do stock markets have predictive content for exchange rate movements?

This paper examines short‐horizon exchange rate predictability and investigates whether stock returns contain information for forecasting daily exchange rate movements. Inspired by the uncovered equity parity condition, we show that stock return differentials have in‐sample and out‐of‐sample predictive power for nominal exchange rates with short horizons (1‐day‐ahead predictions). That is, stock markets inform us about exchange rate movements, at least in the case of high‐frequency data.     

Cerivastatin: a cellular and molecular drug for the future?

The 'statin story' began in 1987 when the first-generation, fungal HMG-CoA reductase inhibitor lovastatin received FDA approval in the USA. Ten years later, the sixth compound of this class came onto the world market - the fully synthetic statin cerivastatin. A number of clinical studies had confirmed its high pharmacological efficacy, its excellent pharmacokinetic properties with fast and nearly complete absorption after oral uptake, a linear kinetic over a broad concentration range, and its favorable safety profile. The greatest advantages, of cerivastatin, however, are its lipophilicity, its high bioavailability of about 60% after oral application and its potency at 100-fold lower doses compared to other lipophilic statins. Nevertheless, the most exciting findings are certainly its non-lipid-related, pleiotropic effects at the cellular and molecular level. Statin therapy was also found to reduce mortality in cases where cholesterol levels or atherosclerotic plaque formation remained unaltered. However, cerivastatin improves endothelial dysfunction, possesses anti-inflammatory, antioxidant, anticoagulant, antithrombotic, antiproliferative, plaque-stabilizing, immunmodulatory, and angiogenic effects, and may even prevent tumor growth, Alzheimer's disease, and osteoporosis. Most of these effects seem to be based on the inhibition of isoprenoid synthesis. Although cerivastatin is no longer on the market because of some problematic side effects, it could be one of the most potent cellular and molecular drugs for the future. Received 29 May 2002; received after revision 23 August 2002; accepted 26 August 2002 RID="*" ID="*"Corresponding author.     

The cucumber tendril — a new test organ for gibberellic acid

Résumé On peut se servir de tronçons de vrilles de concombre en voie de dévelopmentin vitro, pour déterminer d'après leurs réactions la présence d'acide gibberellique (GA). L'avantage de cette méthode réside dans le fait qu'on peut l'employer en la combinant avec la chromatographie sur papier et ainsi évaluer le GA dans l'extrait des plantes.     

Leukocyte recruitment in atherosclerosis: Potential targets for therapeutic approaches?

Atherosclerosis is a complex inflammatory disease involving cellular migration and interaction. Vascular injury in response to different cardiovascular risk factors enhances endothelial dysfunction, which in turn promotes the expression of inflammatory markers and transendothelial leukocyte migration. Recruitment of leukocytes from the blood stream into the vessel intima is a crucial step for the development of the disease. Recent findings have highlighted the role of chemokines, chemokine receptors, adhesion molecules, and gap junctions in this process by acting as chemoattractant, adhesive, or intercellular communication molecules. In this short review, we summarize new data concerning the different steps from leukocyte arrest to transendothelial migration and discuss potential new therapeutic approaches concerning these processes. Received 15 March 2006; received after revision 19 May 2006; accepted 13 June 2006