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Dynamic Nanoproteins: Self-Assembly of Peptides on Monolayer Protected Gold Nanoparticles
Full text linkProtein-protein interactions mediate a large number of important regulatory pathways in the organism and also play a central role in many pathologies. Protein surface recognition provides a powerful tool for the regulation of those protein-protein interactions. Gold nanoparticles offer a suitable platform for multi-functionalization with a wide range of biological ligands for the selective binding and detection of biological targets such as proteins. In particular, gold nanoparticles functionalized with peptide fragments are very attractive for that purpose, since peptides contain all necessary chemical information to interact with natural proteins. During the past years, self-assembly, i.e. the spontaneous organization of molecules into ordered aggregates, has emerged as the most attractive way to develop highly complex nanosized systems. Au NPs containing 1,4,7-triazacyclononane (TACN)·Zn(II) head groups (Au NP 1) have been shown to be attractive scaffolds for the formation of multivalent supramolecular structures.
In this PhD-project the self-assembly of small peptides on monolayer protected gold nanoparticles has been studied as a means to develop dynamic nanoproteins for application in biomolecular recognition and catalysis. The first part has been dedicated to the identification and optimization of small peptides able to bind with high affinity to the surface of Au NP 1, which are gold nanoparticles (d ∼ 2 nm) covered with a monolayer of C9-thiols containing a 1,4,7-triazacyclonane•Zn(II) head group. The results showed four potential candidates of which the tripeptide LWS(p) (S(p) = phosphoserine) had the highest affinity. From a series of studies in which the metal-ion in the monolayer was varied, it emerged that Zn(II) gave the best results. Using LWS(p) as a lead structure a small peptide library was synthesized successively in which additional amino acid residues were attached to the binding unit. Amino acids containing negatively or positively charged, polar and apolar side chains were chosen in order to create a chemical diverse library. Subsequent binding studies showed that all peptides had a very high affinity for Au NP 1, apart from the peptides containing positively charged amino acids.
Subsequently, the peptide library was used to self-assembly a dynamic nanoprotein by adding all peptides simultaneously to Au NP 1. Binding studies revealed that binding occurred under saturation conditions at low micromolar concentrations in aqueous buffer at pH = 7. The addition of a competitor for binding resulted in a complete displacement of the peptides demonstrating the dynamic nature of the surface. In this way it has been demonstrated that it is possible to build up a complex multivalent system in a straightforward manner.
The dynamic nature of the system was exploited in a series of self-selection experiments, aimed at determining how the surface composition on Au NP 1 would change when an excess of peptides would be present. A new protocol relying on the use of ultracentrifugation filters containing a MW cut-off membrane was developed for the purpose of analyzing the surface composition. The results showed a spontaneous self-selection of the peptides with a higher affinity for Au NP 1 upon increasing the overall concentration of the peptide library.
In the next stage, the ability of the nanoprotein to interact with natural protein surfaces was investigated. A particular attention was paid to exploitation of the dynamic nature of the assembly. The serine protease chymotrypsin (ChT) was chosen as a target, because previous literature reports had already mentioned that peptide-functionalized Au NPs are able to bind ChT. Binding assays confirmed that the presence of ChT did not affect the interaction between the peptide library and Au NP 1. Apart from peptide P2, none of the peptides was hydrolyzed by ChT. Enzyme activity studies in the presence of Au NP 1-peptide systems did not provide conclusive data. The obtained data showed a slight activation of ChT (1.5 times) in the presence of Au NP, irrespective of whether peptides were present or not. Such an activation of ChT by cationic agents has also been reported in other studies, but a clear explanation is not yet available. Since the measurement of enzyme activity is an indirect method of measuring the interaction between the nanoproteins and ChT, the attention was shifted towards a direct method relying on the ultracentrifugation experiments developed earlier. The aim was to investigate whether the surface composition would be affected by the addition of ChT. Initial data from the ultracentrifugation experiments and additional fluorescence studies seemed to suggest the formation of a ternary multivalent complex with peptide P1 sandwiched between Au NP 1 and ChT. However, further experiments under different conditions and including other techniques (such as ITC and DLS) are required to confirm these results.
Finally, a collaboration with the Ulijn group at the University of Strathclyde led to a detailed investigation of the activity of Au NP 1 as artificial phosphatases. The original idea was to exploit the high affinity of peptides for Au NP 1 to shift the equilibrium of a dynamic system of interconverting peptides (catalyzed by thermolysin). However, initial studies immediately revealed that after some days the nanoparticles-enzyme mixture caused the dephosphorylation of Fmoc-Yp-OH. This observation led to an in depth study of the origin of this reaction. Initial experiments performed at high concentrations pointed to thermolysin as the prime responsible for the dephosphorylation reaction assisted by Zn(II) or Au NP 1. Under these conditions hardly any activity by Au NP 1 was observed. However, the picture changed completely when the kinetic studies were repeated at much lower (micro)molar concentrations. Now, a strong catalytic activity of Au NP 1 was detected. Indeed, at conditions, i.e. Fmoc-Yp-OH closer to the SSC of Au NP 1, only 10 hours were needed to convert 50% of substrate. Although for a precise understanding of the mechanism more studies are required, the fact that Au NP 1 is able to cause the dephosphorylation of a monophosphate is a very exciting result, considering that the catalytic activity of Au NP 1 and related systems has so far mainly been limited to activated phosphodiesters
Dynamic nanoproteins: Self-assembled peptide surfaces on monolayer protected gold nanoparticles
No full textHere, we demonstrate the formation of dynamic peptide surfaces through the self-assembly of small peptides on the surface of monolayer protected gold nanoparticles. The complexity of the peptide surface can be simply tuned by changing the chemical nature of the added peptides and the ratio in which these are added. The dynamic nature of the surface permits adaptation to changes in the environment
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
Author-wise bibliometric analysis based on entropy.
No full textAuthor-wise bibliometric analysis based on entropy.</p
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