754 research outputs found
Three-dimensional fluorescent microscopy via simultaneous illumination and detection at multiple planes.
The conventional optical microscope is an inherently two-dimensional (2D) imaging tool. The objective lens, eyepiece and image sensor are all designed to capture light emitted from a 2D 'object plane'. Existing technologies, such as confocal or light sheet fluorescence microscopy have to utilize mechanical scanning, a time-multiplexing process, to capture a 3D image. In this paper, we present a 3D optical microscopy method based upon simultaneously illuminating and detecting multiple focal planes. This is implemented by adding two diffractive optical elements to modify the illumination and detection optics. We demonstrate that the image quality of this technique is comparable to conventional light sheet fluorescent microscopy with the advantage of the simultaneous imaging of multiple axial planes and reduced number of scans required to image the whole sample volume
Dynamics of allosteric transitions in GroEL
The chaperonin GroEL-GroES, a machine which helps some proteins to fold,
cycles through a number of allosteric states, the state, with high affinity
for substrate proteins (SPs), the ATP-bound state, and the
() complex. Structures are known for each
of these states. Here, we use a self-organized polymer (SOP) model for the
GroEL allosteric states and a general structure-based technique to simulate the
dynamics of allosteric transitions in two subunits of GroEL and the heptamer.
The transition, in which the apical domains undergo counter-clockwise
motion, is mediated by a multiple salt-bridge switch mechanism, in which a
series of salt-bridges break and form. The initial event in the transition, during which GroEL rotates clockwise, involves a
spectacular outside-in movement of helices K and L that results in K80-D359
salt-bridge formation. In both the transitions there is considerable
heterogeneity in the transition pathways. The transition state ensembles (TSEs)
connecting the , , and states are broad with the the
TSE for the transition being more plastic than the TSE. The results suggest that GroEL functions as a
force-transmitting device in which forces of about (5-30) pN may act on the SP
during the reaction cycle.Comment: 32 pages, 10 figures (Longer version than the one published
Optimization and evaluation of a coarse-grained model of protein motion using X-ray crystal data
Simple coarse-grained models, such as the Gaussian Network Model, have been
shown to capture some of the features of equilibrium protein dynamics. We
extend this model by using atomic contacts to define residue interactions and
introducing more than one interaction parameter between residues. We use
B-factors from 98 ultra-high resolution X-ray crystal structures to optimize
the interaction parameters. The average correlation between GNM fluctuation
predictions and the B-factors is 0.64 for the data set, consistent with a
previous large-scale study. By separating residue interactions into covalent
and noncovalent, we achieve an average correlation of 0.74, and addition of
ligands and cofactors further improves the correlation to 0.75. However,
further separating the noncovalent interactions into nonpolar, polar, and mixed
yields no significant improvement. The addition of simple chemical information
results in better prediction quality without increasing the size of the
coarse-grained model.Comment: 18 pages, 4 figures, 1 supplemental file (cnm_si.tex
Markov propagation of allosteric effects in biomolecular systems: application to GroEL–GroES
We introduce a novel approach for elucidating the potential pathways of allosteric communication in biomolecular systems. The methodology, based on Markov propagation of ‘information' across the structure, permits us to partition the network of interactions into soft clusters distinguished by their coherent stochastics. Probabilistic participation of residues in these clusters defines the communication patterns inherent to the network architecture. Application to bacterial chaperonin complex GroEL–GroES, an allostery-driven structure, identifies residues engaged in intra- and inter-subunit communication, including those acting as hubs and messengers. A number of residues are distinguished by their high potentials to transmit allosteric signals, including Pro33 and Thr90 at the nucleotide-binding site and Glu461 and Arg197 mediating inter- and intra-ring communication, respectively. We propose two most likely pathways of signal transmission, between nucleotide- and GroES-binding sites across the cis and trans rings, which involve several conserved residues. A striking observation is the opposite direction of information flow within cis and trans rings, consistent with negative inter-ring cooperativity. Comparison with collective modes deduced from normal mode analysis reveals the propensity of global hinge regions to act as messengers in the transmission of allosteric signals
Time Under the Curve: Assessing the Impact of Regional Lead Treatment Center Home Visit on the Length of Exposure in Lead Poisoned Children
A high proportion of Connecticut residents are at risk of health effects due to lead exposure, especially children under the age of six. The Yale Regional Lead Treatment Center (YRLTC) assists families of children with blood lead levels (BLL) exceeding the Centers for Disease Control and Prevention (CDC) maximum BLL of 5 µg/dL. YRLTC incorporates home visits of lead-exposed children living in Southern Connecticut to mitigate lead poisoning by emphasizing public health initiatives and social work, rather than using a clinic-only approach. This project aimed to evaluate the merits of this new interdisciplinary approach and its tangible effects on health outcomes. Key informant interviews provided perspectives on the value and themes of home visits. Questions focused on physiological measures of lead poisoning and cognitive development, patient interactions during home visits as well as legal, and logistical challenges to lead abatement. The team also shadowed home visits in order to understand the intervention.https://elischolar.library.yale.edu/ysph_pbchrr/1029/thumbnail.jp
oGNM: online computation of structural dynamics using the Gaussian Network Model
An assessment of the equilibrium dynamics of biomolecular systems, and in particular their most cooperative fluctuations accessible under native state conditions, is a first step towards understanding molecular mechanisms relevant to biological function. We present a web-based system, oGNM that enables users to calculate online the shape and dispersion of normal modes of motion for proteins, oligonucleotides and their complexes, or associated biological units, using the Gaussian Network Model (GNM). Computations with the new engine are 5–6 orders of magnitude faster than those using conventional normal mode analyses. Two cases studies illustrate the utility of oGNM. The first shows that the thermal fluctuations predicted for 1250 non-homologous proteins correlate well with X-ray crystallographic data over a broad range [7.3–15 Å] of inter-residue interaction cutoff distances and the correlations improve with increasing observation temperatures. The second study, focused on 64 oligonucleotides and oligonucleotide–protein complexes, shows that good agreement with experiments is achieved by representing each nucleotide by three GNM nodes (as opposed to one-node-per-residue in proteins) along with uniform interaction ranges for all components of the complexes. These results open the way to a rapid assessment of the dynamics of DNA/RNA-containing complexes. The server can be accessed at
Bacteriophages in bathing wàters: A feasibility study on the development of a method based on bacteriophages for the determination of microbiological quality of bathing waters
Projecte: Project report. BCR Information. EU project KINA19506ENC_001. EUROPEAN COMMISSION. Community Research. DG XII/C - Competitive and Sustainable Growth Programme. Published by EU Directorate General ΧΠ - Science, Research and Development ISBN 92-828-9145-3Informe final projecte europeu aigües de bany i bacteriòfagsMethods for the detection and enumeration of somatic coliphages, F-specific RNA bacteriophages and bacteriophages infecting Bacteroides fragilis had been standardised and validated. Conditions for the preparation, transport and distribution of bacteriophage reference materials and preservation of samples had been defined. A method based on flocculation with Mg(OH2) with concentration efficiencies from about 40% was settled to concentrate phages from bathing waters. All methods were successfully implemented in routine laboratories all around the EU. Data on the occurrence of bacteriophages as compared to E. coli and Enterococci are available from diverse situations encountered in the EU. Results allow to conclude that the potential of phages for the determination of the microbiological quality of bathing waters merits to be considered since their determination is feasible and their behaviour in natural water differs from the behaviour of bacterial indicators and consequently they add valuable information
Specificity of Trypsin and Chymotrypsin: Loop Motion Controlled Dynamic Correlation as a Determinant
Trypsin and chymotrypsin are both serine proteases with high sequence and
structural similarities, but with different substrate specificity. Previous
experiments have demonstrated the critical role of the two loops outside the
binding pocket in controlling the specificity of the two enzymes. To understand
the mechanism of such a control of specificity by distant loops, we have used
the Gaussian Network Model to study the dynamic properties of trypsin and
chymotrypsin and the roles played by the two loops. A clustering method was
introduced to analyze the correlated motions of residues. We have found that
trypsin and chymotrypsin have distinct dynamic signatures in the two loop
regions which are in turn highly correlated with motions of certain residues in
the binding pockets. Interestingly, replacing the two loops of trypsin with
those of chymotrypsin changes the motion style of trypsin to chymotrypsin-like,
whereas the same experimental replacement was shown necessary to make trypsin
have chymotrypsin's enzyme specificity and activity. These results suggest that
the cooperative motions of the two loops and the substrate-binding sites
contribute to the activity and substrate specificity of trypsin and
chymotrypsin.Comment: 41 pages, 7 figure
Screened non-bonded interactions in native proteins manipulate optimal paths for robust residue communication
A protein structure is represented as a network of residues whereby edges are
determined by intra-molecular contacts. We introduce inhomogeneity into these
networks by assigning each edge a weight that is determined by amino-acid pair
potentials. Two methodologies are utilized to calculate the average path
lengths (APLs) between pairs: To minimize (i) the maximum weight in the strong
APL, and (ii) the total weight in the weak APL. We systematically screen edges
that have higher than a cutoff potential and calculate the shortest APLs in
these reduced networks, while keeping chain connectivity. Therefore,
perturbations introduced at a selected region of the residue network propagate
to remote regions only along the non-screened edges that retain their ability
to disseminate the perturbation. The shortest APLs computed from the reduced
homogeneous networks with only the strongest few non-bonded pairs closely
reproduce the strong APLs from the weighted networks. The rate of change in the
APL in the reduced residue network as compared to its randomly connected
counterpart remains constant until a lower bound. Upon further link removal,
this property shows an abrupt increase, towards a random coil behavior. Under
different perturbation scenarios, diverse optimal paths emerge for robust
residue communication.Comment: 21 pages with 6 figure
Efficient and Stable Routing Algorithm Based on User Mobility and Node Density in Urban Vehicular Network
Vehicular ad hoc networks (VANETs) are considered an emerging technology in the industrial and educational fields. This technology is essential in the deployment of the intelligent transportation system, which is targeted to improve safety and efficiency of traffic. The implementation of VANETs can be effectively executed by transmitting data among vehicles with the use of multiple hops. However, the intrinsic characteristics of VANETs, such as its dynamic network topology and intermittent connectivity, limit data delivery. One particular challenge of this network is the possibility that the contributing node may only remain in the network for a limited time. Hence, to prevent data loss from that node, the information must reach the destination node via multi-hop routing techniques. An appropriate, efficient, and stable routing algorithm must be developed for various VANET applications to address the issues of dynamic topology and intermittent connectivity. Therefore, this paper proposes a novel routing algorithm called efficient and stable routing algorithm based on user mobility and node density (ESRA-MD). The proposed algorithm can adapt to significant changes that may occur in the urban vehicular environment. This algorithm works by selecting an optimal route on the basis of hop count and link duration for delivering data from source to destination, thereby satisfying various quality of service considerations. The validity of the proposed algorithm is investigated by its comparison with ARP-QD protocol, which works on the mechanism of optimal route finding in VANETs in urban environments. Simulation results reveal that the proposed ESRA-MD algorithm shows remarkable improvement in terms of delivery ratio, delivery delay, and communication overhead
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