Crystal structure determination and dynamic studies of Mycobacterium tuberculosis Cytidine deaminase in complex with products

Cytidine deaminase (CDA) is a key enzyme in the pyrimidine salvage pathway. It is involved in the hydrolytic deamination of cytidine or 2′-deoxycytidine to uridine or 2′-deoxyuridine, respectively. Here we report the crystal structures of Mycobacterium tuberculosis CDA (MtCDA) in complex with uridine (2.4 Å resolution) and deoxyuridine (1.9 Å resolution). Molecular dynamics (MD) simulation was performed to analyze the physically relevant motions involved in the protein–ligand recognition process, showing that structural flexibility of some protein regions are important to product binding. In addition, MD simulations allowed the analysis of the stability of tetrameric MtCDA structure. These findings open-up the possibility to use MtCDA as a target in future studies aiming to the rational design of new inhibitor of MtCDA-catalyzed chemical reaction with potential anti-proliferative activity on cell growth of M. tuberculosis, the major causative agent of tuberculosis.     

The challenges and scope of theoretical biology

Scientific theories seek to provide simple explanations for significant empirical regularities based on fundamental physical and mechanistic constraints. Biological theories have rarely reached a level of generality and predictive power comparable to physical theories. This discrepancy is explained through a combination of frozen accidents, environmental heterogeneity, and widespread non-linearities observed in adaptive processes. At the same time, model building has proven to be very successful when it comes to explaining and predicting the behavior of particular biological systems. In this respect biology resembles alternative model-rich frameworks, such as economics and engineering. In this paper we explore the prospects for general theories in biology, and suggest that these take inspiration not only from physics, but also from the information sciences. Future theoretical biology is likely to represent a hybrid of parsimonious reasoning and algorithmic or rule-based explanation. An open question is whether these new frameworks will remain transparent to human reason. In this context, we discuss the role of machine learning in the early stages of scientific discovery. We argue that evolutionary history is not only a source of uncertainty, but also provides the basis, through conserved traits, for very general explanations for biological regularities, and the prospect of unified theories of life.     

Cytophotometric determination of the binding of basic dyes by DNA following acetylation

Summary DNA was removed from various tissues by histochemical acetylation of amino groups in proteins using pure acetic anhydride, as demonstrated by cytophotometric (UV, Feulgen, gallocyanin chromalum) and biochemical techniques. Since new phosphate groups were simultaneously exposed, the intensity of methylene blue staining was increased in spite of the nucleic acid release. Under conditions where no extraction occurs the staining intensity increases for more than 30 per cent. On the other hand, the staining intensity of gallocyanin chromalum kept constant. As it had been demonstrated previously, that gallocyanin chromalum binds to about 86 per cent of the DNA phosphate groups, it was concluded that this dye binds to a higher percentage of phosphate groups than do the usual basic dyes. Since it is not possible under the conditions used to make all nucleic acid phosphate groups available for basic dye binding by blocking the amino groups of proteins it can be assumed that not only electrostatic, but also spatial and steric relationships influence the binding capacity of basic dyes to the phosphate groups of nucleoproteins.Supported by a grant from the Deutsche Forschungsgemeinschaft, Bad Godesberg, Germany.     

The effect of excess moisture on the germination of Spinacia oleracea L.

Summary The reversible inhibition of the germination of spinach (Spinacia oleracea L.) seeds in conditions which are even slightly wetter than optimal has been traced to the production, in a wet environment, of a layer of mucilage around and within the fruit coat which surrounds the true seed. Such wet seeds may however germinate readily when the temperature is lowered, or the oxygen pressure of the environment is raised, or the intact seeds are placed for a short time in hydrogen peroxide before being transferred to what normally would be an excess of water. Even in the absence of an increased oxygen supply the seeds will germinate under water provided the fruit coat, or even a small part of it where it covers the radicle, is crefully removed. No evidence has been found of a water soluble inhibitor and the findings are consistent with the hypothesis that germination is dependent on a sufficiently high rate of supply of oxygen to the sites embryonic respiration. The mucilage which is formed under wet conditions forms a barrier which prevents the transfer of oxygen to the embryo by gaseous diffusion or aqueous convection currents and restricts it to the process of aqueous diffusion, and under these conditions the rate of oxygen supply may not reach the threshold level required for germination.     

Creative Commons licenses and the non-commercial condition: Implications for the re-use of biodiversity information

The Creative Commons (CC) licenses are a suite of copyright-based licenses defining terms for the distribution and re-use of creative works. CC provides licenses for different use cases and includes open content licenses such as the Attribution license (CC BY, used by many Open Access scientific publishers) and the Attribution Share Alike license (CC BY-SA, used by Wikipedia, for example). However, the license suite also contains non-free and non-open licenses like those containing a "non-commercial" (NC) condition. Although many people identify "non-commercial" with "non-profit", detailed analysis reveals that significant differences exist and that the license may impose some unexpected re-use limitations on works thus licensed. After providing background information on the concepts of Creative Commons licenses in general, this contribution focuses on the NC condition, its advantages, disadvantages and appropriate scope. Specifically, it contributes material towards a risk analysis for potential re-users of NC-licensed works.     

Switches, excitable responses and oscillations in the Ring1B/Bmi1 ubiquitination system

In an active, self-ubiquitinated state, the Ring1B ligase monoubiquitinates histone H2A playing a critical role in Polycomb-mediated gene silencing. Following ubiquitination by external ligases, Ring1B is targeted for proteosomal degradation. Using biochemical data and computational modeling, we show that the Ring1B ligase can exhibit abrupt switches, overshoot transitions and self-perpetuating oscillations between its distinct ubiquitination and activity states. These different Ring1B states display canonical or multiply branched, atypical polyubiquitin chains and involve association with the Polycomb-group protein Bmi1. Bistable switches and oscillations may lead to all-or-none histone H2A monoubiquitination rates and result in discrete periods of gene (in)activity. Switches, overshoots and oscillations in Ring1B catalytic activity and proteosomal degradation are controlled by the abundances of Bmi1 and Ring1B, and the activities and abundances of external ligases and deubiquitinases, such as E6-AP and USP7.     

Cholinergic innervation of the retrosplenial cortex via the fornix pathway as determined by high affinity choline uptake,choline acetyltransferase activity,and muscarinic receptor binding in the rat

The cholinergic projections from basal forebrain nuclei to the retrosplenial cortex (RSC) have previously been studied using a variety of histological approaches. Studies using acetylcholinesterase (AChE) histochemistry and choline acetyltransferase (ChAT) immunocytochemistry have demonstrated that this projection travels via the cingulum on route to the RSC. Preliminary studies from our laboratory, however, have shown that the fornix may also be involved in this projection. The present study uses the combination of pathway lesions, and the analysis of cholinergic neurochemical markers in the RSC to determine the role of the fornix in the cholinergic projection to the RSC. High affinity choline uptake (HACU) and ChAT activity were measured in the RSC of control rats, animals with cingulate lesions, and animals with fornix plus cingulate lesions. Fornix plus cingulate lesions resulted in significant deceases in HACU and ChAT activity in comparison to cingulate lesions alone. Muscarinic receptor binding was also evaluated in combination with the various lesions, and a significant increase in retrosplenial receptor binding was noted following fornix lesions. Together, these results support the concept of a fornix-mediated cholinergic pathway to the RSC.     

Induction of accessory cell function of human alveolar macrophages by inhalation of human natural interleukin-2

 Accessory function allows antigen-presenting cells to produce sufficient secondary signals for optimum T cell proliferation and interleukin-2 (IL-2) production. Alveolar macrophages are inferior accessory cells compared to monocytes (PBM). We report here that the accessory index (AI) of alveolar macrophages and PBM of patients with lung metastases of solid tumors treated with inhalations of human natural IL-2 (hnIL-2) increased following its administration (P<0.005). The accessory index was significantly elevated from baseline values after 2 weeks of inhalation of 300 000 IU hnIL-2/day (8.2±10.2 compared to 1.1±1; P<0.001). The inhalation of 150 000 IU also induced increases in the index (AI = 2.3±1.9), however, without reaching statistical significance. In addition at 300 000 IU IL-2/day a significant increase in the accessory index was observed for PBM (4±2.5; P<0.05). The indices of PBM and alveolar macrophages prior to inhalation showed a significant negative correlation with the age of the patients (r _s =  – 0.5; r _s =  – 0.8, respectively; P<0.03 for all comparisons). Our data demonstrate that the inhalational application of hnIL-2 enhances the accessory function of alveolar macrophages and, to lesser extent, the accessory index of PBM, indicating the occurrence of pharmacological immunostimulation. Received: 16 August 1995 / Accepted: 4 January 1996