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Cardiovascular interventional MR imaging: a new road for therapy and repair in the heart.

Over the last 5 years, interventional MR imaging has been fertile ground for research. Real-time MR imaging, combined with recent advances in other MR imaging modalities such as perfusion imaging and intravascular imaging, has opened up new paths for cardiac therapy. The recent reports on cardiac stem cell therapy guided and monitored by MR imaging suggest that we are already seeing the establishment of an important role for cardiac MR imaging in cardiac restoration. The collaborative effort from a multidisciplinary team of basic biologists, engineers, and clinicians will refine stem cell incubation and labeling for MR-guided transcatheter endomyocardial injections, and this in turn may facilitate new studies in humans. Several groups have demonstrated in animal studies the feasibility of MR-guided catheter interventions for the treatment of congenital heart disease and arrythmia therapy. Hence, applications in humans remain the challenge for the next years. Although there have been first reports of cardiac catheterizations in humans by combined use of x-ray fluoroscopy and MR imaging, there are no reports in the literature suggesting that active tracking methods by MR imaging have been applied to humans. Safety issues (namely, heating of catheters and wires) hamper clinical use, particularly in infants and children. Current reports are promising that these limitations will be overcome in the near future and will eventually reduce x-ray usage during catheterization. In its current state, cardiac MR imaging offers a unique opportunity to investigate new therapeutic strategies for the treatment of congenital and acquired heart disease.

Cardiac Catheterization↗

Topography for independent binding of alpha-helical and PPII-helical ligands to a peroxisomal SH3 domain.

While the function of most small signaling domains is confined to binary ligand interactions, the peroxisomal Pex13p SH3 domain has the unique capacity of binding to two different ligands, Pex5p and Pex14p. We have used this domain as a model to decipher its structurally independent ligand binding sites. By the combined use of X-ray crystallography, NMR spectroscopy, and circular dichroism, we show that the two ligands bind in unrelated conformations to patches located at opposite surfaces of this SH3 domain. Mutations in the Pex13p SH3 domain that abolish interactions within the Pex13p-Pex5p interface specifically impair PTS1-dependent protein import into yeast peroxisomes.

Amino Acid Motifs↗

Enhancement of spin coherence using Q-factor engineering in semiconductor microdisc lasers.

Semiconductor microcavities offer unique means of controlling light-matter interactions in confined geometries, resulting in a wide range of applications in optical communications and inspiring proposals for quantum information processing and computational schemes. Studies of spin dynamics in microcavities, a new and promising research field, have revealed effects such as polarization beats, stimulated spin scattering and giant Faraday rotation. Here, we study the electron spin dynamics in optically pumped GaAs microdisc lasers with quantum wells and interface-fluctuation quantum dots in the active region. In particular, we examine how the electron spin dynamics are modified by the stimulated emission in the discs, and observe an enhancement of the spin-coherence time when the optical excitation is in resonance with a high-quality (Q approximately 5,000) lasing mode. This resonant enhancement, contrary to expectations from the observed trend in the carrier-recombination time, is then manipulated by altering the cavity design and dimensions. In analogy with devices based on excitonic coherence, this ability to engineer coherent interactions between electron spins and photons may provide new pathways towards spin-dependent quantum optoelectronics.

Biomedical Engineering↗

Determination of helix-helix interactions in membranes by rotational resonance NMR.

Dimerization of human glycophorin A in erythrocyte membranes is mediated by specific interactions within the helical transmembrane domain of the protein. Rotational resonance NMR provides a unique approach for obtaining high-resolution structural data in membrane systems and has been used to establish intermolecular contacts in the glycophorin A dimer by using hydrophobic peptides that correspond to the transmembrane sequence. Magnetization exchange rates were measured between [13C]methyl labels in the hydrophobic sequence -G79-V80-M81-A82-G83-V84- located in the middle of the transmembrane domain and specific [13C]carbonyl labels along the peptide backbone across the dimer interface. Significant magnetization exchange was observed only between V80 (13CH3) and G79 (13C = O) and between V84 (13CH3) and G83 (13C = O), indicating that these residues are packed in the dimer interface in a "ridges-ingrooves" arrangement.

Amino Acid Sequence↗

Crystal structure of Mycobacterium tuberculosis D-3-phosphoglycerate dehydrogenase: extreme asymmetry in a tetramer of identical subunits.

Phosphoglycerate dehydrogenases exist in at least three different structural motifs. The first D-3-phosphoglycerate dehydrogenase structure to be determined was from Escherichia coli and is a tetramer composed of identical subunits that contain three discernable structural domains. The crystal structure of D-3-phosphoglycerate dehydrogenase from Mycobacterium tuberculosis has been determined at 2.3 A. This enzyme represents a second structural motif of the D-3-phosphoglycerate dehydrogenase family, one that contains an extended C-terminal region. This structure is also a tetramer of identical subunits, and the extended motif of 135 amino acids exists as a fourth structural domain. This intervening domain exerts quite a surprising characteristic to the structure by introducing significant asymmetry in the tetramer. The asymmetric unit is composed of two identical subunits that exist in two different conformations characterized by rotation of approximately 180 degrees around a hinge connecting two of the four domains. This asymmetric arrangement results in the formation of two different and distinct domain interfaces between identical domains in the asymmetric unit. As a result, the surface of the intervening domain that is exposed to solvent in one subunit is turned inward in the other subunit toward the center of the structure where it makes contact with other structural elements. Significant asymmetry is also seen at the subunit level where different conformations exist at the NAD-binding site and the putative serine-binding site in the two unique subunits.

Amino Acid Motifs↗

IELs: enforcing law and order in the court of the intestinal epithelium.

The intestinal intraepithelial lymphocytes (IELs) are mostly T cells dispersed as single cells within the epithelial cell layer that surrounds the intestinal lumen. IELs are, therefore, strategically located at the interface between the antigen-rich outside world and the sterile core of the body. The intestine of higher vertebrates has further evolved to harbor numerous commensal bacteria that carry out important functions for the host, and while defensive immunity can effectively protect against the invasion of pathogens, similar immune reactions against food-derived antigens or harmless colonizing bacteria can result in unnecessary and sometimes damaging immune responses. Probably as a result of this unique dilemma imposed by the gut environment, multiple subsets of IEL have differentiated, which all display characteristics of 'activated yet resting' immune cells. Despite this common feature, IELs are heterogeneous with regard to their phenotype, ontogeny, and function. In this review, we discuss the different subtypes of IELs and highlight the distinct pathways they took that led to their unique differentiation into highly specialized effector memory T cells, which provide the most effective immune protection yet in a strictly regulated fashion to preserve the integrity and vital functions of the intestinal mucosal epithelium.

Animals↗

A body model server for human motion capture and representation.

This paper presents a body model server (BMS) that provides real-time access to the position and posture of a person's torso, arms, hands, head, and eyes. It can be accessed by clients over a network. The BMS is designed to function as a device-independent data-layer between the sensing devices and client applications that require real-time human motion data, such as animation control. It can provide clients with accurate information at up to 40 Hz. For data collection, the model uses four magnetic position/orientation sensors, two data-gloves, and an eye-tracker. The BMS combines the data-streams from the sensors and transforms them into snapshots of the user's upper-body pose. A geometric model made up of joints and segments structures the input. Posture of the body is represented by joint angles. Two unique characteristics of our approach are the use of the implicit, geometric constraints of the sensed body to simplify the computation of the unmeasured joint angles, and the use of time-stamped data that allow synchronization with other data streams, e.g., speech input. This paper describes the architecture of the BMS, including the management of multiple input devices, the representation and computation of the position and joint angle data, and the client-server interface.

Computer Simulation↗

Spatial ability subfactors and their influences on a computer-based information search task.

OBJECTIVE: The present study examined the relationship between two distinct subfactors of spatial ability and performance in an information search task modeled on browsing the Web. BACKGROUND: Previous studies have found relationships between various measures of spatial ability and performance in a wide variety of computer-based tasks. METHOD: In the search task 101 participants (18-29 years of age) searched for the answer to a question by navigating the system. They completed the experimental task as well as a battery of cognitive ability measures that included two different measures of spatial ability. RESULTS: The results indicate that spatial orientation ability was related to performance with tasks that were high in their navigational requirement (engendered by the use of a novel aid), whereas spatial visualization was unrelated to performance in any task condition. CONCLUSION: A closer inspection of the cognitive requirements of a task may reveal what interventions could be most useful when designing computer systems or developing training programs. APPLICATION: Given the unique differences between the different spatial abilities, the current results suggest the design of navigational aids that place less demand on spatial orientation ability.

Adolescent↗

Orientation distributions for cytochrome c on polar and nonpolar interfaces by total internal reflection fluorescence.

The formation of chemisorbed monolayers of yeast cytochrome c on both uncharged polar and nonpolar soft surfaces of organic self-assembled monolayers (SAM) on solid inorganic substrates was followed in situ by polarized total internal reflection fluorescence. Two types of nonpolar surfaces and one type of uncharged polar surface were used. The first type of nonpolar surface contained only thiol endgroups, while the other was composed of a mixture of thiol and methyl endgroups. The uncharged polar surface was provided by the mixture of thiol and hydroxyl endgroups. The thiol endgroups were used to form a covalent disulfide bond with the unique surface-exposed cysteine residue 102 of the protein. The mean tilt angle of the protein's zinc-substituted porphyrin was found to be 41 degrees and 50 degrees for the adsorption onto the nonpolar and uncharged polar surfaces, respectively. The distribution widths for the pure thiol and the thiol/methyl and thiol/hydroxyl mixtures were 9 degrees, 1 degrees, and 18 degrees, respectively. The high degree of the orientational order and good stability achieved for the protein monolayer on the mixed thiol/methyl endgroup SAM makes this system very attractive for studies of both intramolecular and intermolecular electron transfer processes.

Biophysical Phenomena↗

Characterization of glycosphingolipids by supercritical fluid chromatography-mass spectrometry.

Gangliosides have been characterized by supercritical fluid chromatography-chemical ionization mass spectrometry (SFC-CIMS) as permethyl and pertrimethylsilyl derivatives, using carbon dioxide as the SFC mobile phase and CI reagent gas. Ganglioside classes and ceramide heterogeneity within each class are well resolved by SFC. Direct SFC-interfacing allows the analytical manipulations of single-ion monitoring, total-ion plots, background subtraction, library searches, and spectral reconstruction algorithms. Addition of ammonia to the CI ion chamber (NH3 as a CI reagent gas) yields abundant molecular-weight-related ions, (MH)+ and (MNH4)+ from analyte derivatives. Substitution of methanol for ammonia yields considerable parent-ion fragmentation, providing structural information on carbohydrate sequence, fatty acid, and sphingoid components. Under these latter conditions a unique alpha-cleavage fragment is observed which differentiates fatty acid from sphingosine heterogeneity. For ganglioside samples, the carboxyl group of neuraminyl residue(s) have been esterified with pentafluorobenzyl bromide and the products analyzed by negative ion chemical ionization MS. This modification improves chemical selectivity and greatly enhances detecting sensitivity. These "soft" ionization conditions provide abundant molecular-weight-related anions for collision-induced dissociation and subpicogram detection.

Ammonia↗

ProMate: a structure based prediction program to identify the location of protein-protein binding sites.

Is the whole protein surface available for interaction with other proteins, or are specific sites pre-assigned according to their biophysical and structural character? And if so, is it possible to predict the location of the binding site from the surface properties? These questions are answered quantitatively by probing the surfaces of proteins using spheres of radius of 10 A on a database (DB) of 57 unique, non-homologous proteins involved in heteromeric, transient protein-protein interactions for which the structures of both the unbound and bound states were determined. In structural terms, we found the binding site to have a preference for beta-sheets and for relatively long non-structured chains, but not for alpha-helices. Chemically, aromatic side-chains show a clear preference for binding sites. While the hydrophobic and polar content of the interface is similar to the rest of the surface, hydrophobic and polar residues tend to cluster in interfaces. In the crystal, the binding site has more bound water molecules surrounding it, and a lower B-factor already in the unbound protein. The same biophysical properties were found to hold for the unbound and bound DBs. All the significant interface properties were combined into ProMate, an interface prediction program. This was followed by an optimization step to choose the best combination of properties, as many of them are correlated. During optimization and prediction, the tested proteins were not used for data collection, to avoid over-fitting. The prediction algorithm is fully automated, and is used to predict the location of potential binding sites on unbound proteins with known structures. The algorithm is able to successfully predict the location of the interface for about 70% of the proteins. The success rate of the predictor was equal whether applied on the unbound DB or on the disjoint bound DB. A prediction is assumed correct if over half of the predicted continuous interface patch is indeed interface. The ability to predict the location of protein-protein interfaces has far reaching implications both towards our understanding of specificity and kinetics of binding, as well as in assisting in the analysis of the proteome.

Algorithms↗

Resolving the fluorescence response of Escherichia coli carbamoyl phosphate synthetase: mapping intra- and intersubunit conformational changes.

Carbamoyl phosphate synthetase (CPS) from Escherichia coli is potentially overlaid with a network of allosterism, interconnecting active sites, effector binding sites, and aggregate interfaces to control its mechanisms of catalytic synchronization, regulation, and oligomerization, respectively. To characterize these conformational changes, a tryptophan-free variant of CPS was genetically engineered by substituting six native tryptophans with tyrosines. Each tryptophan was then reinserted, singly, as a specific fluorescence probe of its corresponding microenvironment. The amino acid substitutions themselves result in little apparent disruption of the protein; variants maintain catalytic and allosteric functionality, and the fluorescence properties of each tryptophan, while unique, are additive to wild-type CPS. Whereas the collective, intrinsic fluorescence response of E. coli CPS is largely insensitive to ligand binding, changes of the individual probes in intensity, lifetime, anisotropy, and accessibility to acrylamide quenching highlight the dynamic interplay between several protein domains, as well as between subunits. W213 within the carboxy phosphate domain, for example, exhibits an almost 40% increase in intensity upon saturation with ATP; W437 of the oligomerization domain, in contrast, is essentially silent in its fluorescence to the binding of ligands. Nucleotide and bicarbonate association within the large subunit induces fluorescence changes in both W170 and W175 of the small subunit, indicative of the type of long-range interactions purportedly synchronizing the carboxy phosphate and amidotransferase domains of the enzyme to initiate catalysis. ATP and ADP engender different fluorescence responses in most tryptophans, perhaps reflecting coordinating, conformational changes accompanying the cycling of reactants and products during catalysis.

Adenosine Triphosphate↗

Crystal structure of yeast cytosine deaminase. Insights into enzyme mechanism and evolution.

Yeast cytosine deaminase is an attractive candidate for anticancer gene therapy because it catalyzes the deamination of the prodrug 5-fluorocytosine to form 5-fluorouracil. We report here the crystal structure of the enzyme in complex with the inhibitor 2-hydroxypyrimidine at 1.6-A resolution. The protein forms a tightly packed dimer with an extensive interface of 1450 A2 per monomer. The inhibitor was converted into a hydrated adduct as a transition-state analog. The essential zinc ion is ligated by the 4-hydroxyl group of the inhibitor together with His62, Cys91, and Cys94 from the protein. The enzyme shares similar active-site architecture to cytidine deaminases and an unusually high structural homology to 5-aminoimidazole-4-carboxamide-ribonucleotide transformylase and thereby may define a new superfamily. The unique C-terminal tail is involved in substrate specificity and also functions as a gate controlling access to the active site. The complex structure reveals a closed conformation, suggesting that substrate binding seals the active-site entrance so that the catalytic groups are sequestered from solvent. A comparison of the crystal structures of the bacterial and fungal cytosine deaminases provides an elegant example of convergent evolution, where starting from unrelated ancestral proteins, the same metal-assisted deamination is achieved through opposite chiral intermediates within distinctly different active sites.

Amino Acid Sequence↗

Inherent protein structural flexibility at the RNA-binding interface of L30e.

The Saccharomyces cerevisiae ribosomal protein L30 autoregulates its own expression by binding to a purine-rich internal loop in its pre-mRNA and mRNA. NMR studies of L30 and its RNA complex showed that both the internal loop of the RNA as well as a region of the protein become substantially more ordered upon binding. A crystal structure of a maltose binding protein (MBP)-L30 fusion protein with two copies in the asymmetric unit has been determined. The flexible RNA-binding region in the L30 copies has two distinct conformations, one resembles the RNA bound form solved by NMR and the other is unique. Structure prediction algorithms also had difficulty accurately predicting this region, which is consistent with conformational flexibility seen in the NMR and X-ray crystallography studies. Inherent conformational flexibility may be a hallmark of regions involved in intermolecular interactions.

Amino Acid Sequence↗

Flexible Structural Neighborhood--a database of protein structural similarities and alignments.

Protein structures are flexible, changing their shapes not only upon substrate binding, but also during evolution as a collective effect of mutations, deletions and insertions. A new generation of protein structure comparison algorithms allows for such flexibility; they go beyond identifying the largest common part between two proteins and find hinge regions and patterns of flexibility in protein families. Here we present a Flexible Structural Neighborhood (FSN), a database of structural neighbors of proteins deposited in PDB as seen by a flexible protein structure alignment program FATCAT, developed previously in our group. The database, searchable by a protein PDB code, provides lists of proteins with statistically significant structural similarity and on lower menu levels provides detailed alignments, interactive superposition of structures and positions of hinges that were identified in the comparison. While superficially similar to other structural protein alignment resources, FSN provides a unique resource to study not only protein structural similarity, but also how protein structures change. FSN is available from a server http://fatcat.burnham.org/fatcat/struct_neighbor and by direct links from the PDB database.

Databases, Protein↗

Remifentanil update: clinical science and utility.

The anilidopiperidine opioid remifentanil has pharmacodynamic properties similar to all opioids; however, its pharmacokinetic characteristics are unique. Favourable pharmacokinetic properties, minimally altered by extremes of age or renal or hepatic dysfunction, enable easy titration and rapid dissipation of clinical effect of this agent, even after prolonged infusion. Remifentanil is metabolised by esterases that are widespread throughout the plasma, red blood cells, and interstitial tissues, whereas other anilidopiperidine opioids (e.g. fentanyl, alfentanil and sufentanil) depend upon hepatic biotransformation and renal excretion for elimination. Consequently, remifentanil is cleared considerably more rapidly than other anilidopiperidine opioids. In addition, its pKa (the pH at which the drug is 50% ionised) is less than physiological pH; thus, remifentanil circulates primarily in the non-ionised moiety, which quickly penetrates the lipid blood-brain barrier and rapidly equilibrates across the plasma/effect site interface. By virtue of these distinctive pharmacokinetic properties, the context-sensitive half-time (i.e. the time required for the drug's plasma concentration to decrease by 50% after cessation of an infusion) of remifentanil remains consistently short (3.2 minutes), even following an infusion of long duration (> or =8 hours). Remifentanil, a clinically versatile opioid, is useful for intravenous analgesia and sedation in spontaneously breathing patients undergoing painful procedures. Profound analgesia may be achieved with minimal effect on cognitive function. Remifentanil may also provide sedation and analgesia during placement of regional anaesthetic blocks, and in conjunction with topical anaesthesia and airway nerve blocks, it may be useful for blunting reflex responses and facilitating 'awake' fibreoptic intubation. Compared with fentanyl and alfentanil in a day-surgery setting, remifentanil supplementation of general anaesthesia may improve intraoperative haemodynamic control. Both emergence time and the incidence of respiratory depression during post-anaesthetic recovery may be reduced. However, outcomes such as home discharge time, post-emergence adverse effect profile, and patient and provider satisfaction are not significantly improved, and the incidence of intraoperative hypotension and bradycardia is greater. In addition, drug acquisition costs for remifentanil are higher and clinicians may need extra time to familiarise themselves with the drug's unique pharmacokinetics.Ironically, the quick dissipation of opioid analgesic effect following remifentanil discontinuation may be a significant clinical disadvantage. Unless little or no postoperative pain is anticipated, the clinician may wish to treat prospectively using local or regional anaesthesia, non-opioid analgesics, or longer-acting opioid analgesics.

Analgesics, Opioid↗

Rapid separation of phenylthiohydantoin amino acids: ambient pressure ion-mobility mass spectrometry (IMMS).

An electrospray ionization (ESI) ambient pressure ion-mobility spectrometer (APIMS) interfaced to an orthogonal reflector time-of-flight mass spectrometer (TOFMS) was evaluated for the first time as a detector for the identification of phenylthiohydantoin (PTH)-derivatized amino acids, the final products in the Edman sequencing process of peptides and proteins. The drift and flight times of the twenty common PTH amino acids were characterized by a well-defined 2-D mobility/mass spectral pattern. The combination of mobility/mass modes of analysis gave rise to a unique trend-line formation for the series of PTH amino acids. In addition, each PTH amino acid had a unique reduced mobility constant K(o), thus enabling the differentiation of all the amino acid derivatives including the PTH-leucine and PTH-isoleucine isomers. More importantly it was shown that it was possible to resolve a complete reference mixture of PTH amino acids in a single experimental run in less than 1 min. Detection limits for the PTH amino acids were found to range from 1.04 to 3.52 ng; indicating that the limits of detection were less than 17.0 pmol for all of the PTH amino acids.

Amino Acids↗

Endothelial cells of the brain and other organ systems: some similarities and differences.

The cerebral endothelium represents an active interface between blood and central nervous system. The blood-brain barrier restricts the free passage of nutrients, hormones, drugs and cellular elements to the brain. Recent studies performed on freshly isolated cerebral microvessels and cultured endothelial cells of brain capillaries provided a better understanding of the properties and functions of cerebral endothelial cells. This review summarizes the main findings of the in vitro approach in the blood-brain barrier research, describes the common endothelial and unique cerebral features of the brain endothelium, and provides a short overview on how these blood-brain barrier characteristics can be induced in cerebral endothelial cells by the neighbouring cells.

Blood-Brain Barrier↗