Efficient Sampling of Protein Loop Regions Using Conformational Hashing Complemented with Random Coordinate Descent.

construction of loop regions is an important problem in computational structural biology. Compared to regions with well-defined secondary structure, loops tend to exhibit significant conformational heterogeneity. As a result, their structures are often ambiguous when determined using experimental data obtained by crystallography, cryo-EM, or NMR.

Specificity of the chromophore-binding site in human cone opsins.

The variable composition of the chromophore-binding pocket in visual receptors is essential for vision. The visual phototransduction starts with the isomerization of the retinal chromophore upon absorption of photons. Despite sharing the common 11--retinal chromophore, rod and cone photoreceptors possess distinct photochemical properties.

Crowd sourcing difficult problems in protein science.

Dedicated computing resources are expensive to develop, maintain, and administrate. Frequently, research groups require bursts of computing power, during which progress is still limited by available computing resources. One way to alleviate this bottleneck would be to use additional computing resources.

Complex binding pathways determine the regeneration of mammalian green cone opsin with a locked retinal analogue.

Phototransduction is initiated when the absorption of light converts the 11--retinal chromophore to its all- configuration in both rod and cone vertebrate photoreceptors. To sustain vision, 11--retinal is continuously regenerated from its all- conformation through a series of enzymatic steps comprising the "visual or retinoid" cycle. Abnormalities in this cycle can compromise vision because of the diminished supply of 11--retinal and the accumulation of toxic, constitutively active opsin.

Hydrogen/Deuterium Exchange Mass Spectrometry of Human Green Opsin Reveals a Conserved Pro-Pro Motif in Extracellular Loop 2 of Monostable Visual G Protein-Coupled Receptors.

Opsins comprise the protein component of light sensitive G protein-coupled receptors (GPCRs) in the retina of the eye that are responsible for the transduction of light into a biochemical signal. Here, we used hydrogen/deuterium (H/D) exchange coupled with mass spectrometry to map conformational changes in green cone opsin upon light activation. We then compared these findings with those reported for rhodopsin.

Structural Insights into the Drosophila melanogaster Retinol Dehydrogenase, a Member of the Short-Chain Dehydrogenase/Reductase Family.

The 11-cis-retinylidene chromophore of visual pigments isomerizes upon interaction with a photon, initiating a downstream cascade of signaling events that ultimately lead to visual perception. 11-cis-Retinylidene is regenerated through enzymatic transformations collectively called the visual cycle. The first and rate-limiting enzymatic reaction within this cycle, i.

Image registration and averaging of low laser power two-photon fluorescence images of mouse retina.

Two-photon fluorescence microscopy (TPM) is now being used routinely to image live cells for extended periods deep within tissues, including the retina and other structures within the eye . However, very low laser power is a requirement to obtain TPM images of the retina safely. Unfortunately, a reduction in laser power also reduces the signal-to-noise ratio of collected images, making it difficult to visualize structural details.

Characterization of the Domain Orientations of E. coli 5'-Nucleotidase by Fitting an Ensemble of Conformers to DEER Distance Distributions.

Escherichia coli 5'-nucleotidase is a two-domain enzyme exhibiting a unique 96° domain motion that is required for catalysis. Here we present an integrated structural biology study that combines DEER distance distributions with structural information from X-ray crystallography and computational biology to describe the population of presumably almost isoenergetic open and closed states in solution. Ensembles of models that best represent the experimental distance distributions are determined by a Monte Carlo search algorithm.

Semi-automated discrimination of retinal pigmented epithelial cells in two-photon fluorescence images of mouse retinas.

Automated image segmentation is a critical step toward achieving a quantitative evaluation of disease states with imaging techniques. Two-photon fluorescence microscopy (TPM) has been employed to visualize the retinal pigmented epithelium (RPE) and provide images indicating the health of the retina. However, segmentation of RPE cells within TPM images is difficult due to small differences in fluorescence intensity between cell borders and cell bodies.

BCL::MP-fold: Membrane protein structure prediction guided by EPR restraints.

For many membrane proteins, the determination of their topology remains a challenge for methods like X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy. Electron paramagnetic resonance (EPR) spectroscopy has evolved as an alternative technique to study structure and dynamics of membrane proteins. The present study demonstrates the feasibility of membrane protein topology determination using limited EPR distance and accessibility measurements.

Postoperative observation of children after endoscopic type 1 posterior laryngeal cleft repair.

Objectives: To report the perioperative management and surgical outcomes in a large series of pediatric patients with endoscopically repaired type 1 posterior laryngeal cleft (PLC).

Study Design: Case series with chart review.

Setting: Urban, tertiary care, free-standing pediatric hospital.

Noninvasive two-photon microscopy imaging of mouse retina and retinal pigment epithelium through the pupil of the eye.

Two-photon excitation microscopy can image retinal molecular processes in vivo. Intrinsically fluorescent retinyl esters in subcellular structures called retinosomes are an integral part of the visual chromophore regeneration pathway. Fluorescent condensation products of all-trans-retinal accumulate in the eye with age and are also associated with age-related macular degeneration (AMD).

Energetic analysis of the rhodopsin-G-protein complex links the α5 helix to GDP release.

We present a model of interaction of Gi protein with the activated receptor (R*) rhodopsin, which pinpoints energetic contributions to activation and reconciles the β2 adrenergic receptor-Gs crystal structure with new and previously published experimental data. In silico analysis demonstrated energetic changes when the Gα C-terminal helix (α5) interacts with the R* cytoplasmic pocket, thus leading to displacement of the helical domain and GDP release. The model features a less dramatic domain opening compared with the crystal structure.

RosettaEPR: rotamer library for spin label structure and dynamics.

An increasingly used parameter in structural biology is the measurement of distances between spin labels bound to a protein. One limitation to these measurements is the unknown position of the spin label relative to the protein backbone. To overcome this drawback, we introduce a rotamer library of the methanethiosulfonate spin label (MTSSL) into the protein modeling program Rosetta.

Pediatric obstructive sleep apnea syndrome.

Pediatric obstructive sleep apnea syndrome (OSAS) is a common health problem diagnosed and managed by various medical specialists, including family practice physicians, pediatricians, pulmonologists, and general and pediatric otolaryngologists. If left untreated, the sequelae can be severe. Over the last decade, significant advancements have been made in the evidence-based management of pediatric OSAS.

Conformation of receptor-bound visual arrestin.

Arrestin-1 (visual arrestin) binds to light-activated phosphorylated rhodopsin (P-Rh*) to terminate G-protein signaling. To map conformational changes upon binding to the receptor, pairs of spin labels were introduced in arrestin-1 and double electron-electron resonance was used to monitor interspin distance changes upon P-Rh* binding. The results indicate that the relative position of the N and C domains remains largely unchanged, contrary to expectations of a "clam-shell" model.

Treatment strategies for lateral sphenoid sinus recess cerebrospinal fluid leaks.

Objective: To highlight concepts critical to achieving successful repair and avoiding intracranial complications in the treatment of cerebrospinal fluid (CSF) leaks from the lateral recess of the sphenoid sinus (LRS).

Design: Outcomes study.

Setting: Tertiary referral university hospital.

Cystic fibrosis transmembrane conductance regulator modulation by the tobacco smoke toxin acrolein.

Objectives/hypothesis: Evidence indicates that decreased mucociliary clearance (MCC) is a major contributing feature to chronic rhinosinusitis. Tobacco-smoke exposure is thought to inhibit transepithelial Cl(-) secretion, a major determinant of airway surface liquid hydration and MCC. The objective of the current study was to evaluate the effects of acrolein exposure (a prominent tobacco smoke toxin) on vectorial Cl(-) transport through the major apical anion channel cystic fibrosis transmembrane conductance regulator (CFTR) in sinonasal epithelium.

MRSA and non-MRSA otorrhea in children: a comparative study of clinical course.

Objective: To test the perception that post-tympanostomy tube otorrhea caused by methicillin-resistant Staphylococcus aureus (MRSA) is a more virulent disease than otorrhea caused by other pathogens by analyzing the clinical differences and disease courses in children diagnosed with otorrhea caused by MRSA bacteria vs non-MRSA bacteria.

Design: Retrospective review.

Setting: Tertiary children's hospital.

Fixation methods in pediatric cochlear implants: retrospective review of an evolution of 3 techniques.

Objective: To review 3 techniques of cochlear implant (CI) fixation used by a single surgeon for the fixation of 320 consecutive CIs in a pediatric population and associated complications.

Study Design: Case series with chart review.

Setting: Tertiary referral children's hospital.

Resveratrol has salutary effects on mucociliary transport and inflammation in sinonasal epithelium.

Objective/hypothesis: Therapeutic agents that enhance mucociliary transport (via stimulation of transepithelial Cl- secretion) and inhibit inflammation could provide considerable advantages over conventional treatments for chronic rhinosinusitis (CRS). The objectives of the present study were to investigate whether the polyphenolic compound resveratrol promotes transepithelial Cl- transport and inhibits KC/IL-8 secretion in sinonasal epithelium.

Study Design: In vitro and in vivo study.

Algorithm for selection of optimized EPR distance restraints for de novo protein structure determination.

A hybrid protein structure determination approach combining sparse Electron Paramagnetic Resonance (EPR) distance restraints and Rosetta de novo protein folding has been previously demonstrated to yield high quality models (Alexander et al. (2008)). However, widespread application of this methodology to proteins of unknown structures is hindered by the lack of a general strategy to place spin label pairs in the primary sequence.

The management of myringotomy tubes in pediatric cochlear implant recipients.

Objective: To investigate the impact of myringotomy tubes (MTs) on outcomes for pediatric cochlear implant (CI) recipients.

Design: Retrospective case-control chart review.

Setting: Tertiary care pediatric hospital.

Transduction of multiple cell types using improved conditions for gene delivery and expression of SV40 pseudovirions packaged in vitro.

This comprehensive study demonstrates highly efficient transduction of a wide variety of human, murine, and monkey cell lines, using a procedure for in vitro packaging of plasmid DNA in recombinant simian virus 40 (SV40) capsid proteins to form pseudovirions. The pseudovirions are encapsidated by the VP1 major capsid protein, with no SV40 sequence requirement, and are able to carry up to 17.7 kb of supercoiled plasmid DNA.