Site-Directed Mutagenesis of the Cysteine Ligands to the [4Fe−4S] Cluster of Escherichia coli MutY †

International audienceThe Escherichia coli DNA repair enzyme MutY plays an important role in the recognition and repair of 7,8-dihydro-8-oxo-2′-deoxyguanosine:2′-deoxyadenosine (OG:A) mismatches in DNA [Michaels et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 7022-7025]. MutY prevents DNA mutations resulting from the misincorporation of A opposite OG by using N-glycosylase activity to remove the adenine base. An interesting feature of MutY is that it contains a [4Fe-4S] 2+ cluster that has been shown to play an important role in substrate recognition [Porello, S. L., Cannon, M. J., David, S. S. (1998) Biochemistry 37, 6465-6475]. Herein, we have used site-directed mutagenesis to individually replace the cysteine ligands to the [4Fe-4S] 2+ cluster of E. coli MutY with serine, histidine, and alanine. The extent to which the various mutations reduce the levels of protein overexpression suggests that coordination of the [4Fe-4S] 2+ cluster provides stability to MutY in vivo. The ability of the mutated enzymes to bind to a substrate analogue DNA duplex and their in vivo activity were evaluated. Remarkably, the effects are both substitution and position dependent. For example, replacement of cysteine 199 with histidine provides a mutated enzyme that is expressed at high levels and exhibits DNA binding and in vivo activity similar to the WT enzyme. These results suggest that histidine coordination to the iron-sulfur cluster may be accommodated at this position in MutY. In contrast, replacement of cysteine 192 with histidine results in less efficient DNA binding and in vivo activity compared to the WT enzyme without affecting levels of overexpression. The results from the site-directed mutagenesis suggest that the structural properties of the iron-sulfur cluster coordination domain are important for both substrate DNA recognition and the in vivo activity of MutY

Collective emotions online and their influence on community life

E-communities, social groups interacting online, have recently become an object of interdisciplinary research. As with face-to-face meetings, Internet exchanges may not only include factual information but also emotional information - how participants feel about the subject discussed or other group members. Emotions are known to be important in affecting interaction partners in offline communication in many ways. Could emotions in Internet exchanges affect others and systematically influence quantitative and qualitative aspects of the trajectory of e-communities? The development of automatic sentiment analysis has made large scale emotion detection and analysis possible using text messages collected from the web. It is not clear if emotions in e-communities primarily derive from individual group members' personalities or if they result from intra-group interactions, and whether they influence group activities. We show the collective character of affective phenomena on a large scale as observed in 4 million posts downloaded from Blogs, Digg and BBC forums. To test whether the emotions of a community member may influence the emotions of others, posts were grouped into clusters of messages with similar emotional valences. The frequency of long clusters was much higher than it would be if emotions occurred at random. Distributions for cluster lengths can be explained by preferential processes because conditional probabilities for consecutive messages grow as a power law with cluster length. For BBC forum threads, average discussion lengths were higher for larger values of absolute average emotional valence in the first ten comments and the average amount of emotion in messages fell during discussions. Our results prove that collective emotional states can be created and modulated via Internet communication and that emotional expressiveness is the fuel that sustains some e-communities.Comment: 23 pages including Supporting Information, accepted to PLoS ON

Inhaled alpha 1 -proteinase inhibitor therapy in patients with cystic fibrosis

Inhaled alpha1-proteinase inhibitor (PI) is known to reduce neutrophil elastase burden in some patients with CF. This phase 2a study was designed to test inhaled Alpha-1 HC, a new aerosolized alpha1-PI formulation, in CF patients

Using a runway paradigm to assess the relative strength of rats' motivations for enrichment objects

Laboratory animals should be provided with enrichment objects in their cages; however, it is first necessary to test whether the proposed enrichment objects provide benefits that increase the animals’ welfare. The two main paradigms currently used to assess proposed enrichment objects are the choice test, which is limited to determining relative frequency of choice, and consumer demand studies, which can indicate the strength of a preference but are complex to design. Here, we propose a third methodology: a runway paradigm, which can be used to assess the strength of an animal’s motivation for enrichment objects, is simpler to use than consumer demand studies, and is faster to complete than typical choice tests. Time spent with objects in a standard choice test was used to rank several enrichment objects in order to compare with the ranking found in our runway paradigm. The rats ran significantly more times, ran faster, and interacted longer with objects with which they had previously spent the most time. It was concluded that this simple methodology is suitable for measuring rats’ motivation to reach enrichment objects. This can be used to assess the preference for different types of enrichment objects or to measure reward system processes

A Fast Algorithm Finding the Shortest Reset Words

In this paper we present a new fast algorithm finding minimal reset words for finite synchronizing automata. The problem is know to be computationally hard, and our algorithm is exponential. Yet, it is faster than the algorithms used so far and it works well in practice. The main idea is to use a bidirectional BFS and radix (Patricia) tries to store and compare resulted subsets. We give both theoretical and practical arguments showing that the branching factor is reduced efficiently. As a practical test we perform an experimental study of the length of the shortest reset word for random automata with $n$ states and 2 input letters. We follow Skvorsov and Tipikin, who have performed such a study using a SAT solver and considering automata up to $n=100$ states. With our algorithm we are able to consider much larger sample of automata with up to $n=300$ states. In particular, we obtain a new more precise estimation of the expected length of the shortest reset word $\approx 2.5\sqrt{n-5}$.Comment: COCOON 2013. The final publication is available at http://link.springer.com/chapter/10.1007%2F978-3-642-38768-5_1

Emotional persistence in online chatting communities

How do users behave in online chatrooms, where they instantaneously read and write posts? We analyzed about 2.5 million posts covering various topics in Internet relay channels, and found that user activity patterns follow known power-law and stretched exponential distributions, indicating that online chat activity is not different from other forms of communication. Analysing the emotional expressions (positive, negative, neutral) of users, we revealed a remarkable persistence both for individual users and channels. I.e. despite their anonymity, users tend to follow social norms in repeated interactions in online chats, which results in a specific emotional "tone" of the channels. We provide an agent-based model of emotional interaction, which recovers qualitatively both the activity patterns in chatrooms and the emotional persistence of users and channels. While our assumptions about agent's emotional expressions are rooted in psychology, the model allows to test different hypothesis regarding their emotional impact in online communication.Comment: 34 pages, 4 main and 12 supplementary figure

Emergence of good conduct, scaling and Zipf laws in human behavioral sequences in an online world

We study behavioral action sequences of players in a massive multiplayer online game. In their virtual life players use eight basic actions which allow them to interact with each other. These actions are communication, trade, establishing or breaking friendships and enmities, attack, and punishment. We measure the probabilities for these actions conditional on previous taken and received actions and find a dramatic increase of negative behavior immediately after receiving negative actions. Similarly, positive behavior is intensified by receiving positive actions. We observe a tendency towards anti-persistence in communication sequences. Classifying actions as positive (good) and negative (bad) allows us to define binary 'world lines' of lives of individuals. Positive and negative actions are persistent and occur in clusters, indicated by large scaling exponents alpha~0.87 of the mean square displacement of the world lines. For all eight action types we find strong signs for high levels of repetitiveness, especially for negative actions. We partition behavioral sequences into segments of length n (behavioral `words' and 'motifs') and study their statistical properties. We find two approximate power laws in the word ranking distribution, one with an exponent of kappa-1 for the ranks up to 100, and another with a lower exponent for higher ranks. The Shannon n-tuple redundancy yields large values and increases in terms of word length, further underscoring the non-trivial statistical properties of behavioral sequences. On the collective, societal level the timeseries of particular actions per day can be understood by a simple mean-reverting log-normal model.Comment: 6 pages, 5 figure

Positive words carry less information than negative words

We show that the frequency of word use is not only determined by the word length \cite{Zipf1935} and the average information content \cite{Piantadosi2011}, but also by its emotional content. We have analyzed three established lexica of affective word usage in English, German, and Spanish, to verify that these lexica have a neutral, unbiased, emotional content. Taking into account the frequency of word usage, we find that words with a positive emotional content are more frequently used. This lends support to Pollyanna hypothesis \cite{Boucher1969} that there should be a positive bias in human expression. We also find that negative words contain more information than positive words, as the informativeness of a word increases uniformly with its valence decrease. Our findings support earlier conjectures about (i) the relation between word frequency and information content, and (ii) the impact of positive emotions on communication and social links.Comment: 16 pages, 3 figures, 3 table

Positivity of the English language

Over the last million years, human language has emerged and evolved as a fundamental instrument of social communication and semiotic representation. People use language in part to convey emotional information, leading to the central and contingent questions: (1) What is the emotional spectrum of natural language? and (2) Are natural languages neutrally, positively, or negatively biased? Here, we report that the human-perceived positivity of over 10,000 of the most frequently used English words exhibits a clear positive bias. More deeply, we characterize and quantify distributions of word positivity for four large and distinct corpora, demonstrating that their form is broadly invariant with respect to frequency of word use.Comment: Manuscript: 9 pages, 3 tables, 5 figures; Supplementary Information: 12 pages, 3 tables, 8 figure

Wetting films on chemically heterogeneous substrates

Based on a microscopic density functional theory we investigate the morphology of thin liquidlike wetting films adsorbed on substrates endowed with well-defined chemical heterogeneities. As paradigmatic cases we focus on a single chemical step and on a single stripe. In view of applications in microfluidics the accuracy of guiding liquids by chemical microchannels is discussed. Finally we give a general prescription of how to investigate theoretically the wetting properties of substrates with arbitrary chemical structures.Comment: 56 pages, RevTeX, 20 Figure