A systematic computational analysis of biosynthetic gene cluster evolution: lessons for engineering biosynthesis.

Bacterial secondary metabolites are widely used as antibiotics, anticancer drugs, insecticides and food additives. Attempts to engineer their biosynthetic gene clusters (BGCs) to produce unnatural metabolites with improved properties are often frustrated by the unpredictability and complexity of the enzymes that synthesize these molecules, suggesting that genetic changes within BGCs are limited by specific constraints. Here, by performing a systematic computational analysis of BGC evolution, we derive evidence for three findings that shed light on the ways in which, despite these constraints, nature successfully invents new molecules: 1) BGCs for complex molecules often evolve through the successive merger of smaller sub-clusters, which function as independent evolutionary entities.

Comparative Protein Structure Modeling Using MODELLER.

Functional characterization of a protein sequence is one of the most frequent problems in biology. This task is usually facilitated by accurate three-dimensional (3-D) structure of the studied protein. In the absence of an experimentally determined structure, comparative or homology modeling can sometimes provide a useful 3-D model for a protein that is related to at least one known protein structure.

Uncertainty in integrative structural modeling.

Integrative structural modeling uses multiple types of input information and proceeds in four stages: (i) gathering information, (ii) designing model representation and converting information into a scoring function, (iii) sampling good-scoring models, and (iv) analyzing models and information. In the first stage, uncertainty originates from data that are sparse, noisy, ambiguous, or derived from heterogeneous samples. In the second stage, uncertainty can originate from a representation that is too coarse for the available information or a scoring function that does not accurately capture the information.

Molecular architecture of the 40S⋅eIF1⋅eIF3 translation initiation complex.

Eukaryotic translation initiation requires the recruitment of the large, multiprotein eIF3 complex to the 40S ribosomal subunit. We present X-ray structures of all major components of the minimal, six-subunit Saccharomyces cerevisiae eIF3 core. These structures, together with electron microscopy reconstructions, cross-linking coupled to mass spectrometry, and integrative structure modeling, allowed us to position and orient all eIF3 components on the 40S⋅eIF1 complex, revealing an extended, modular arrangement of eIF3 subunits.

Structural characterization by cross-linking reveals the detailed architecture of a coatomer-related heptameric module from the nuclear pore complex.

Most cellular processes are orchestrated by macromolecular complexes. However, structural elucidation of these endogenous complexes can be challenging because they frequently contain large numbers of proteins, are compositionally and morphologically heterogeneous, can be dynamic, and are often of low abundance in the cell. Here, we present a strategy for the structural characterization of such complexes that has at its center chemical cross-linking with mass spectrometric readout.

Molecular architecture of photoreceptor phosphodiesterase elucidated by chemical cross-linking and integrative modeling.

Photoreceptor phosphodiesterase (PDE6) is the central effector enzyme in visual excitation pathway in rod and cone photoreceptors. Its tight regulation is essential for the speed, sensitivity, recovery and adaptation of visual detection. Although major steps in the PDE6 activation/deactivation pathway have been identified, mechanistic understanding of PDE6 regulation is limited by the lack of knowledge about the molecular organization of the PDE6 holoenzyme (αβγγ).

Integrative structure-function mapping of the nucleoporin Nup133 suggests a conserved mechanism for membrane anchoring of the nuclear pore complex.

The nuclear pore complex (NPC) is the sole passageway for the transport of macromolecules across the nuclear envelope. Nup133, a major component in the essential Y-shaped Nup84 complex, is a large scaffold protein of the NPC's outer ring structure. Here, we describe an integrative modeling approach that produces atomic models for multiple states of Saccharomyces cerevisiae (Sc) Nup133, based on the crystal structures of the sequence segments and their homologs, including the related Vanderwaltozyma polyspora (Vp) Nup133 residues 55 to 502 (VpNup133(55-502)) determined in this study, small angle X-ray scattering profiles for 18 constructs of ScNup133 and one construct of VpNup133, and 23 negative-stain electron microscopy class averages of ScNup133(2-1157).

Determining protein complex structures based on a Bayesian model of in vivo Förster resonance energy transfer (FRET) data.

The use of in vivo Förster resonance energy transfer (FRET) data to determine the molecular architecture of a protein complex in living cells is challenging due to data sparseness, sample heterogeneity, signal contributions from multiple donors and acceptors, unequal fluorophore brightness, photobleaching, flexibility of the linker connecting the fluorophore to the tagged protein, and spectral cross-talk. We addressed these challenges by using a Bayesian approach that produces the posterior probability of a model, given the input data. The posterior probability is defined as a function of the dependence of our FRET metric FRETR on a structure (forward model), a model of noise in the data, as well as prior information about the structure, relative populations of distinct states in the sample, forward model parameters, and data noise.

Cys-scanning disulfide crosslinking and bayesian modeling probe the transmembrane signaling mechanism of the histidine kinase, PhoQ.

Bacteria transduce signals across the membrane using two-component systems (TCSs), consisting of a membrane-spanning sensor histidine kinase and a cytoplasmic response regulator. In gram-negative bacteria, the PhoPQ TCS senses cations and antimicrobial peptides, yet little is known about the structural changes involved in transmembrane signaling. We construct a model of PhoQ signal transduction using Bayesian inference, based on disulfide crosslinking data and homologous crystal structures.

Molecular architecture and function of the SEA complex, a modulator of the TORC1 pathway.

The TORC1 signaling pathway plays a major role in the control of cell growth and response to stress. Here we demonstrate that the SEA complex physically interacts with TORC1 and is an important regulator of its activity. During nitrogen starvation, deletions of SEA complex components lead to Tor1 kinase delocalization, defects in autophagy, and vacuolar fragmentation.

Insights into secondary metabolism from a global analysis of prokaryotic biosynthetic gene clusters.

Although biosynthetic gene clusters (BGCs) have been discovered for hundreds of bacterial metabolites, our knowledge of their diversity remains limited. Here, we used a novel algorithm to systematically identify BGCs in the extensive extant microbial sequencing data. Network analysis of the predicted BGCs revealed large gene cluster families, the vast majority uncharacterized.

The role of reward prediction in the control of attention.

Previously rewarded stimuli involuntarily capture attention. The learning mechanisms underlying this value-driven attentional capture remain less understood. We tested whether theories of prediction-based associative reward learning explain the conditions under which reward feedback leads to value-based modulations of attentional priority.

Randomized controlled trial examining the effects of fish oil and multivitamin supplementation on the incorporation of n-3 and n-6 fatty acids into red blood cells.

The present randomized, placebo-controlled, double-blind, parallel-groups clinical trial examined the effects of fish oil and multivitamin supplementation on the incorporation of n-3 and n-6 fatty acids into red blood cells. Healthy adult humans (n = 160) were randomized to receive 6 g of fish oil, 6 g of fish oil plus a multivitamin, 3 g of fish oil plus a multivitamin or a placebo daily for 16 weeks. Treatment with 6 g of fish oil, with or without a daily multivitamin, led to higher eicosapentaenoic acid (EPA) composition at endpoint.

Prediction of substrates for glutathione transferases by covalent docking.

Enzymes in the glutathione transferase (GST) superfamily catalyze the conjugation of glutathione (GSH) to electrophilic substrates. As a consequence they are involved in a number of key biological processes, including protection of cells against chemical damage, steroid and prostaglandin biosynthesis, tyrosine catabolism, and cell apoptosis. Although virtual screening has been used widely to discover substrates by docking potential noncovalent ligands into active site clefts of enzymes, docking has been rarely constrained by a covalent bond between the enzyme and ligand.

Elucidating the mechanism of substrate recognition by the bacterial Hsp90 molecular chaperone.

Hsp90 is a conformationally dynamic molecular chaperone known to promote the folding and activation of a broad array of protein substrates ("clients"). Hsp90 is believed to preferentially interact with partially folded substrates, and it has been hypothesized that the chaperone can significantly alter substrate structure as a mechanism to alter the substrate functional state. However, critically testing the mechanism of substrate recognition and remodeling by Hsp90 has been challenging.

Can ginger ameliorate chemotherapy-induced nausea? Protocol of a randomized double blind, placebo-controlled trial.

Background: Preliminary research shows ginger may be an effective adjuvant treatment for chemotherapy-induced nausea and vomiting but significant limitations need to be addressed before recommendations for clinical practice can be made.

Methods/design: In a double-blinded randomised-controlled trial, chemotherapy-naïve patients will be randomly allocated to receive either 1.2 g of a standardised ginger extract or placebo per day.

Protein structure modeling with MODELLER.

Genome sequencing projects have resulted in a rapid increase in the number of known protein sequences. In contrast, only about one-hundredth of these sequences have been characterized at atomic resolution using experimental structure determination methods. Computational protein structure modeling techniques have the potential to bridge this sequence-structure gap.

Effects of multivitamin, mineral and herbal supplement on cognition in younger adults and the contribution of B group vitamins.

Objective: Cognitive benefits of multivitamins have been observed in the elderly, but fewer trials have investigated younger, healthy cohorts. This randomised, double-blind, placebo-controlled study investigated the cognitive effects of 16-week multivitamin supplementation in adults aged 20-49 years.

Method: A total of 138 participants aged 20-50 years were randomised and 116 completed the trial.

Orthosis reduces breast pain and mechanical forces through natural and augmented breast tissue in women lying prone.

Background: Breast implant displacement or rupture can cause aesthetic problems and serious medical complications. Activities with prone positioning and loading of the anterior chest wall, such as massage, chiropractic or osteopathic therapies may increase the risk of implant failure and can also cause discomfort in women with natural breast tissue. Here we test the effectiveness of a newly developed orthosis on pain, mechanical pressure and displacement of breast tissue in women with cosmetic augmentation, post-mastectomy reconstruction, lactating or natural breast tissue.

Reinforcement learning modulates the stability of cognitive control settings for object selection.

Cognitive flexibility reflects both a trait that reliably differs between individuals and a state that can fluctuate moment-to-moment. Whether individuals can undergo persistent changes in cognitive flexibility as a result of reward learning is less understood. Here, we investigated whether reinforcing a periodic shift in an object selection strategy can make an individual more prone to switch strategies in a subsequent unrelated task.

SAXS Merge: an automated statistical method to merge SAXS profiles using Gaussian processes.

Small-angle X-ray scattering (SAXS) is an experimental technique that allows structural information on biomolecules in solution to be gathered. High-quality SAXS profiles have typically been obtained by manual merging of scattering profiles from different concentrations and exposure times. This procedure is very subjective and results vary from user to user.

Hepatocyte-like cells derived from human amniotic epithelial cells can be encapsulated without loss of viability or function in vitro.

Placenta derived human amniotic epithelial cells (hAEC) are an attractive source of stem cells for the generation of hepatocyte-like cells (HLC) for therapeutic applications to treat liver diseases. During hAEC differentiation into HLC, they become increasingly immunogenic, which may result in immune cell-mediated rejection upon transplantation into allogeneic recipients. Placing cells within devices such as alginate microcapsules can prevent immune cell-mediated rejection.

The effects of multivitamin supplementation on diurnal cortisol secretion and perceived stress.

Recent evidence suggests that dietary intake of vitamins, in particular the B-vitamins including B6, B9 and B12 may have a number of positive effects on mood and stress. Given the effects of stress on a range of biological mechanisms including the endocrine system, it could be reasonably expected that multivitamin supplementation may also affect markers of these mechanisms such as diurnal cortisol secretion. In the current double-blind placebo-controlled study 138 adults (aged 20 to 50 years) were administered a multivitamin containing B-vitamins versus placebo over a 16-week period.

ModBase, a database of annotated comparative protein structure models and associated resources.

ModBase (http://salilab.org/modbase) is a database of annotated comparative protein structure models. The models are calculated by ModPipe, an automated modeling pipeline that relies primarily on Modeller for fold assignment, sequence-structure alignment, model building and model assessment (http://salilab.