SEARCH · PubMed Health

Results for “Unique interface”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 217 records · Page 12Linked to original sources

PubMed1993

Human nonpancreatic secreted phospholipase A2: interfacial parameters, substrate specificities, and competitive inhibitors.

The rate and equilibrium parameters for the interfacial catalysis by recombinant human nonpancreatic secreted phospholipase A2 were determined. Results show that the enzyme binds to anionic interfaces with considerably higher affinity than to zwitterionic interfaces. The extent of hydrolysis per enzyme on anionic vesicles in the processive scooting mode shows that the enzyme is fully catalytically active as a monomer. Among several secreted phospholipases A2 tested, the human nonpancreatic secreted enzyme is unique in its ability to undergo slow intervesicle exchange either by dissociation from the interface followed by binding to a different vesicle or by promoting the fusion of vesicles. The equilibrium dissociation constants for calcium, substrate analogs, reaction products, and several competitive inhibitors bound to the enzyme at the interface were determined by monitoring the ligand-conferred protection of the active site histidine residue from alkylation by phenacyl bromide. The interfacial Michaelis-Menten parameters were determined from the analysis of the entire reaction progress curve and also by monitoring the effect of competitive inhibitors on the initial rate of hydrolysis in the scooting mode. The interfacial Michaelis constant (KM*) for the substrate 1,2-dimyristoylglycero-sn-3-phosphomethanol was determined to be considerably above the maximal attainable mole fraction of unity for the substrate in the bilayer. Substrate specificity studies show that the enzyme does not significantly discriminate between phospholipids that differ in the type of polar head group or in the degree of unsaturation of the fatty acyl chains. Competitive inhibitors are described that display a high degree of selectivity for binding to the nonpancreatic versus pancreatic phospholipase A2. The kinetic properties of the human nonpancreatic secreted phospholipase A2 suggest that the enzyme has evolved to hydrolyze substrates at anionic interfaces and at high calcium concentrations.

Catalysis↗

PubMed2001

Solution conformation of alpha-conotoxin EI, a neuromuscular toxin specific for the alpha 1/delta subunit interface of torpedo nicotinic acetylcholine receptor.

A high resolution structure of alpha-conotoxin EI has been determined by (1)H NMR spectroscopy and molecular modeling. alpha-Conotoxin EI has the same disulfide framework as alpha 4/7 conotoxins targeting neuronal nicotinic acetylcholine receptors but antagonizes the neuromuscular receptor as do the alpha 3/5 and alpha A conotoxins. The unique binding preference of alpha-conotoxin EI to the alpha(1)/delta subunit interface of Torpedo neuromuscular receptor makes it a valuable structural template for superposition of various alpha-conotoxins possessing distinct receptor subtype specificities. Structural comparison of alpha-conotoxin EI with the gamma-subunit favoring alpha-conotoxin GI suggests that the Torpedo delta-subunit preference of the former originates from its second loop. Superposition of three-dimensional structures of seven alpha-conotoxins reveals that the estimated size of the toxin-binding pocket in nicotinic acetylcholine receptor is approximately 20 A (height) x 20 A (width) x 15 A (thickness).

Amino Acid Sequence↗

PubMed2000

Scanning tunneling microscopy: a unique tool in the study of chirality, dynamics, and reactivity in physisorbed organic monolayers.

Scanning tunneling microscopy (STM) is applied to study organic monolayers, physisorbed at the liquid-graphite interface. Due to the very local nature of the probing, the structure of these adlayers has been imaged with very high detail. The high resolution allowed us to investigate the effect of molecular chirality on the monolayer formation and provided a unique way to study chemical reactions at the liquid-graphite interface. Making use of a fast scanning mode, dynamic processes in these adlayers have been visualized.

Absorption↗

PubMed1995

A novel low oxygen affinity recombinant hemoglobin (alpha96val--> Trp): switching quaternary structure without changing the ligation state.

Using our Escherichia coli expression plasmid (pHE2) in which synthetic human alpha and beta-globin genes are coexpressed with the E. coli methionine aminopeptidase gene under the control of separate tac promoters, we have constructed a new artificial hemoglobin in which the valine residue at position 96 of the alpha chain, located in the alpha 1 beta 2 subunit interface, has been replaced by a tryptophan residue using site-directed mutagenesis. We have determined the oxygen-binding properties of this recombinant hemoglobin, r Hb (alpha 96Val-->Trp), and have used proton nuclear magnetic resonance spectroscopy to investigate its tertiary structure around the heme group and the quaternary structure in the alpha 1 beta 2 subunit interface. This artificial hemoglobin shows a low oxygen affinity, but high cooperativity in oxygen binding, and exhibits no unusual subunit dissociation when ligated. Molecular dynamics simulations suggest that the unique oxygen-binding property of r Hb (alpha 96Val-->Trp) may be due to an extra hydrogen bond between alpha 96Trp and beta 99Asp in the alpha 1 beta 2 subunit interface in the deoxy form. Despite the replacement of a small amino acid residue, valine, by a large tryptophan residue in the alpha 1 beta 2 subunit interface, this artificial hemoglobin shows very similar tertiary structure around the heme pockets and quaternary structure in the alpha 1 beta 2 subunit interface compared to those of human normal adult hemoglobin. Another unique feature of this artificial hemoglobin is that the ligated form, e.g. carbonmonoxy form, of this hemoglobin in the oxy-quaternary structure can be converted to the deoxy-like quaternary structure by the addition of an allosteric effector, inositol hexaphosphate, as well as by lowering the temperature in the absence of inositol hexaphosphate, without changing its ligation state. Thus, this recombinant hemoglobin can be used to gain new insights regarding the nature of subunit interactions in the alpha 1 beta 2 interface and the molecular basis for the allosteric mechanism of hemoglobin.

Base Sequence↗

PubMed2002

Conformational change coupling the dimerization and activation of KSHV protease.

The mechanism of herpesviral protease activation upon dimerization was studied using two independent spectroscopic assays augmented by directed mutagenesis. Spectroscopic changes, attributable to dimer interface conformational plasticity, were observed upon dimerization of Kaposi's sarcoma-associated herpesvirus protease (KSHV Pr). KSHV Pr's dissociation constant of 585 +/- 135 nM at 37 degrees C was measured by a concentration-dependent, 100-fold increase in specific activity to a value of 0.275 +/- 0.023 microM product min(-1) (microM enzyme)(-1). A 4 nm blue-shifted fluorescence emission spectrum and a 25% increase in ellipticity at 222 nm were detected by circular dichroism upon dimer association. This suggested enhanced hydrophobic packing within the dimer interface and/or core, as well as altered secondary structures. To better understand the structure-activity relationship between the monomer and the dimer, KSHV Pr molecules were engineered to remain monomeric via substitution of two separate residues within the dimer interface, L196 and M197. These mutants were proteolytically inactive while exhibiting the spectroscopic signature and thermal stability of wild type, dissociated monomers (T(M) = 75 degrees C). KSHV Pr conformational changes were found to be relevant in vivo, as the autoproteolytic inactivation of KSHV Pr at its dimer disruption site [Pray et al. (1999) J. Mol. Biol. 289, 197-203] was detected in viral particles from KSHV-infected cells. This characterization of structural plasticity suggests that the structure of the KSHV Pr monomer is stable and significantly different from its structure in the dimer. This structural uniqueness should be considered in the development of compounds targeting the dimer interface of KSHV Pr monomers.

Cell Line↗

PubMed1994

Optimizing recording capabilities of the Utah Intracortical Electrode Array.

The Utah Intracortical Electrode Array is a unique silicon-based monolithic structure designed for use as a multichannel interface to the central nervous system. In this paper, we describe a series of acute experiments designed to determine the neural recording capabilities of this electrode array and the dependence of the signal-to-noise ratio (SNR) of the recordings on the electrode surface area (length of metallized tip). We found that both separable unit and multiunit cluster responses could be recorded. Additionally, high SNR recordings could be achieved for some electrodes (with electrode tip lengths of 30-220 microns), while recordings with signals substantially greater than the noise could be made from most of the electrodes provided that the proper electrode surface area was used. The demonstrated recording capabilities of the Utah Intracortical Electrode Array and its unique three-dimensional structure should form the basis for innovative physiological investigations into the functional organization of the cortex as well as for long term neuroprosthesis development.

Algorithms↗

PubMed1987

The preparative isolation of mitochondria from Chinese hamster ovary cells.

A "hybrid" discontinuous gradient consisting of 6% Percoll overlaid on metrizamide separated mitochondria from other organelles in a Chinese hamster ovary cell postnuclear supernatant in a single 15-min centrifugation. The mitochondrial preparation contained about 25% of the mitochondrial marker, cytochrome-c oxidase, in a form that was about 90% latent. Based on the postnuclear supernatant, cytochrome-c oxidase activity was enriched approximately 45-fold. Trace amounts of lysosomal, rough endoplasmic reticular, Golgi, peroxisomal, plasma membrane, and cytosolic markers were found in the preparation. Electron microscopy revealed that the preparation consisted almost exclusively of mitochondria with only minor amounts of contaminating organelles. Analysis of the mitochondrial preparation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated that the mitochondrial preparation had a unique protein profile compared to the postnuclear supernatant and other gradient interfaces. Separation of the mitochondria into membrane and lumenal (matrix) fractions by treatment with 100 mM Na2CO3, pH 11.5, also indicated that the mitochondria were intact; they were rich in lumenal proteins. The data indicate that the mitochondria represent maximally about 2.2% of Chinese hamster ovary cell postnuclear supernatant protein. These isolated mitochondria should prove useful for problems in molecular cell biology.

Animals↗

PubMed1994

Structure of human chorionic gonadotropin at 2.6 A resolution from MAD analysis of the selenomethionyl protein.

BACKGROUND: Human chorionic gonadotropin (hCG) is a placental hormone that stimulates secretion of the pregnancy-sustaining steroid progesterone. It is a member of a family of glycoprotein hormones that are disulfide-rich heterodimers, with a common alpha-chain and distinctive beta-chains specific to their particular G-protein linked receptors. RESULTS: We have produced recombinant hCG in mammalian cells as the selenomethionyl protein, and have determined its structure (after partial deglycosylation) at 2.6 A resolution from multiwavelength anomalous diffraction (MAD) measurements. Despite only limited sequence similarity (10% identity), the alpha- and beta-subunits of hCG have similar tertiary folds. Each subunit has a cystine-knot motif at its core of extended hairpin loops. There is a very extensive subunit interface featuring two inter-chain beta-sheets and a unique, disulfide-tethered 'arm' from the beta-subunit which 'embraces' the alpha-subunit. The carboxy-terminal peptide of the beta-subunit, which is rich in O-linked sugars, is disordered. CONCLUSIONS: Structural and sequence comparisons indicate an evolutionary homology, albeit remote, between the glycoprotein hormone chains and other cystine-knot proteins, notably platelet-derived growth factor. Segments of the alpha- and beta-chains that have been convincingly implicated in receptor binding by hCG are juxtaposed on one side of the molecule. A glycosylation site implicated in signal transduction but not in binding is also close to the presumed binding site suggesting a possible coupling between ligand binding and signaling. This study with selenomethionyl protein produced in mammalian cells extends the realm of MAD phasing.

Amino Acid Sequence↗

PubMed2002

Complementation of buried lysine and surface polar residues in a designed heterodimeric coiled coil.

The coiled coil is an attractive target for protein design. The helices of coiled coils are characterized by a heptad repeat of residues denoted a to g. Residues at positions a and d form the interhelical interface and are usually hydrophobic. An established strategy to confer structural uniqueness to two-stranded coiled coils is the use of buried polar Asn residues at position a, which imparts dimerization and conformational specificity at the expense of stability. Here we show that polar interactions involving buried position-a Lys residues that can interact favorably only with surface e' or g' Glu residues also impart structural uniqueness to a designed heterodimeric coiled coil with the nativelike properties of sigmoidal thermal and urea-induced unfolding transitions, slow hydrogen exchange and lack of ANS binding. The position-a Lys residues do not, however, confer a single preference for helix orientation, likely reflecting the ability of Lys at position a to from favorable interactions with g' or e' Glu residues in the parallel and antiparallel orientations, respectively. The Lys-Glu polar interaction is less destabilizing than the Asn-Asn a-->a' interaction, presumably reflecting a higher desolvation penalty associated with the completely buried polar position-a groups. Our results extend the range of approaches for two-stranded coiled-coil design and illustrate the role of complementing polar groups associated with buried and surface positions of proteins in protein folding and design.

Amino Acid Motifs↗

PubMed2002

Prostaglandin D2 and reproduction.

This review highlights recent studies investigating the role of prostaglandin (PG)D2 in reproduction. PGD2 induces sleep, allergic responses, inhibition of platelet aggregation, and relaxation of vascular and non-vascular smooth muscle, and has some roles in reproduction. Two types of PGD2 synthase are known. Lipocalin-type PGD synthase is present in cerebrospinal fluid, seminal plasma and may play an important role in male reproduction. Another PGD synthase, hematopoietic PGD synthase is present in the spleen, fallopian tube, endometrial gland cells, extravillous trophoblasts and villous trophoblasts, and perhaps plays an important role in female reproduction. Recent studies demonstrate that PGD2 is probably involved in multiple aspects of inflammation through its dual receptor systems, DP and CRTH2. CRTH2 but not DP is a chemo-attractant receptor for PGD2. Interestingly, CRTH2 is a most reliable marker for the detection of human T helper type 2 (Th2) and T cytotoxic type 2 (Tc2) cells, and the percentages of CRTH expressing CD4+-T cells and CD8+-T cells were significantly higher in the decidua especially at the implantation site, suggesting that Th2 and Tc2 cells recruit into the materno-fetal interface, in a PGD2-mediated manner. PGD2 has a very unique effect to inhibit antigen presentation by inhibition of dendritic cell (DC) migration through DP but not CRTH2. PGD2 might appear to contribute to the maintenance of pregnancy by controlling the Th1/Th2 balance and antigen presentation by DCs through its dual receptor systems, CRTH2 and DP.

Animals↗

PubMed1993

[Diagnosis of rupture of fetal membranes: comparison of vaginal detection of fetal fibronectin and intra-amnion injection of indigo carmine].

Amniotic fluid contains a heavily glycosylated pregnancy-related isoform of fibronectin (fFN), which can be detected using FDC-6, a monoclonal antibody directed against a novel epitope specific to fFN. It has been identified in the amniotic fluid in various concentrations throughout the different pregnancy periods and is localised at the interface of the chorion and the uterine decidua. Because of the unique pregnancy-associated expression of fFN, we determined the utility of cervicovaginal appearance of fFN as a marker of membrane integrity or rupture of membranes. In a prospective study we compared 1) the detection of fFN in cervicovaginal samples by a membrane-immunoassay (ROM-Check, Mast Diagnostica, Reinfeld, FRG) with 2) the results of intraamniotic injection of Indigo Carmine (IC), followed by the observation of dye spill into the vagina to confirm rupture of membranes. 65 patients were examined, 12 patients had to be excluded from the study for various reasons. Examination results were identical in 50 patients, while in three cases the fFN results were positive with a simultaneous negative result of the intraamniotic dye injection. In the case of blood contamination, the fFN test results were not valid; testing of maternal blood and of urine samples was fFN negative in all cases. We conclude, that oncofoetal fibronectin is an excellent marker for rupture of membranes. The test is easy to perform and does not involve the risk of an intraamniotic injection of dye.(ABSTRACT TRUNCATED AT 250 WORDS)

Amnion↗

PubMed1979

Child psychiatry education for general residents.

There is continuing debate about what child psychiatry experiences should be included in a general residency. The author describes the program at the University of Michigan in an effort to provide some insights into the interface between child psychiatry and general residency training. This program is unique in several respects: a 12-month rotation in child psychiatry is offered, and the faculty size and budget of the youth services are comparable to those of the adult services. A survey of all residents and faculty pointed up numerous disagreements as to the length of the rotation and priorities in curriculum. The author discusses the influence of the various competitive processes on the educational program.

Adolescent↗

PubMed1996

Control of skeletal muscle blood flow during dynamic exercise: contribution of endothelium-derived nitric oxide.

Traditional explanations for the hyperaemia which accompanies exercise have invoked the 'metabolic theory' of vasodilation, whereby contractile activity in the active muscle gives rise to metabolic by-products which dilate vessels bathed in interstitial fluid. Whilst metabolites with vasodilator properties have been identified, this theory does not adequately explain the magnitude of hyperaemia observed in active skeletal muscle, principally because large increases in flow are dependent on dilation of 'feed' arteries which lie outside the tissue parenchyma and are not subjected to changes in the interstitial milieu. Coordinated resistance vessel dilation during exercise is therefore dependent on a signal which 'ascends' from the microvessels to the feed arteries located upstream. Recent studies of ascending vasodilation have concentrated on the possible contribution of the endothelium, a monolayer of flattened squamous cells which lie at the interface between the circulating blood and vascular wall. These cells are uniquely positioned to respond to changes in rheological and humoral conditions within the cardiovascular system, and to transduce these changes into vasoactive signals which regulate blood flow, vascular tone and arterial pressure. Endothelial cells produce nitric oxide (NO), a rapidly diffusing labile substance which relaxes adjacent vascular smooth muscle. NO is released basally and contributes to the regulation of vascular tone by acting as a functional antagonist to sympathetic neural constriction. In addition, NO is spontaneously released in response to deformation of the endothelial cell membrane, indicating that changes in pulsatile flow and wall shear stress are likely physiological stimuli. Since the dilation of microvessels in response to exercise increases blood flow through the upstream feed arteries, which subsequently dilate, one explanation for ascending vasodilation is that NO release is stimulated by flow-induced shear stress. Evidence that NO contributes to ascending vasodilation is reviewed, along with studies which indicate that NO mediates exercise hyperaemia, that physical conditioning upregulates NO production and that NO controls blood flow by modifying other physiological mechanisms.

Animals↗

PubMed1976

Resonance ultrasonic measurements of microscopic gas bubbles.

The positive identification and location of stationary or moving bubbles in human tissue is a persisting problem in the study and management of dysbarism. Since bubbles are resonant scatterers of ultrasound, while structures such as red-cell clumps and tissue interfaces are not, it is possible in principle to identify bubbles uniquely and to determine their position and size. A pulsed, ultrasonic echo-ranging system was expressly designed to exploit the principle of bubble resonance, and was evaluated for bubble identification and location in two experiments. Echos from bubbles in a water bath were computer-processed to reveal a distinct "line-narrowing" in the Fourier domain, which is diagnostic of gas bubbles. An increase in echo signal from a dog's jugular vein upon distal bubble injecton was obtained as evidence for in vivo bubble detection. We conclude that the exploitation of the bubble resonance principle can offer a safe, noninvasive technology for micro-bubble identification, location, and size determination in many tissues. This technology appears capable of measuring stationary bubbles in tissue spaces or impacted in blood vessels as well as those moving in the vascular system.

Air↗

PubMed1988

Striatal dopamine and the interface between orienting and ingestive functions.

Recent lesion experiments have implicated forebrain catecholaminergic projections in a unique switching mechanism that enables sensory orientation to occur during ongoing feeding behavior. Unit recording studies indicate that there is a population of neurons in the striatum that respond to tactile stimulation only while an animal is eating. These sensory-related cells may serve as part of a system that diverts attention away from ingestive behavior. In the present study, 6-hydroxydopamine was infused directly into the striatum in rats to eliminate the dopaminergic input specifically to this region. The animals were tested for their reactions to tactile stimulation of the vibrissae during or in the absence of eating. During noneating trials, orienting was rapid and reliable to stimuli presented on either side of the body midline. While eating, contralateral orienting never occurred, even when the stimulation was intense, whereas ipsilateral orienting was unaffected. It was suggested that the capacity to disengage from ingestive behavior may depend importantly on the integrity of dopaminergic input to the striatum.

Animals↗

PubMed1984

Structural comparison of the two distinct sugar binding sites in wheat germ agglutinin isolectin II.

The two unique sugar binding sites in wheat germ agglutinin, located in the subunit/subunit interface of the dimer molecule and termed primary and secondary binding sites, are compared in the light of the newly obtained chemical amino acid sequence and a high-resolution electron density map (1.8 A). Homology was found in the three amino acid residues directly involved in sugar binding: Tyr73II, Ser62II, Glu115I in the primary site, and Tyr159I, Ser148I, Asp29II in the secondary site (subscripts refer to promoters I and II). Thirteen corresponding side-chain atoms of these three homologous residues in the two sites could be superimposed with a root-mean-square difference of 1.39 A. The three sugar binding residues are located in subsite 1 of each extended binding location and contribute to binding of the terminal, non-reducing N-acetyl-D-glucosamine and N-acetyl-D-neuraminic acid residues only, and they provide three hydrogen bonds for complex stabilization. Two hydrogen bonds are made with the carbonyl and amido portions of the N-acetyl group and the third with the C-3 OH group of the sugar ring. It is suggested that small differences in the sugar binding affinities at these two unique sites exist, due to the different numbers of van der Waals' interactions made at these sites, which contribute to stabilizing, for instance, the wheat germ agglutinin/N,N'-diacetyl-chitobiose complex. The single tryptophan residue is located at a distance of approximately 13 A from the primary site and is thought to have no affect on sugar binding. In addition, the disposition of the four saccharide binding sites of the dimer with respect to three local, pseudo 2-fold symmetry axes, relating domains of opposite protomers, is discussed.

Amino Acid Sequence↗

PubMed1991

Coordinate developmental regulation of purine catabolic enzyme expression in gastrointestinal and postimplantation reproductive tracts.

Using histochemical detection, we have visualized in situ the complete metabolic pathway for the degradation of purine nucleotides. From the tongue to the ileum, diverse epithelial cell types lining the lumen of the mouse gastrointestinal (GI) tract strongly coexpress each of the five key purine catabolic enzymes. Dramatic increases in the expression of each enzyme occurred during postnatal maturation of the GI tract. Using in situ hybridization, an intense accumulation of adenosine deaminase (ADA) mRNA was detected only within GI epithelial cells undergoing postmitotic differentiation. In a similar manner, at the developing maternal-fetal interface, high level expression of the purine catabolic pathway also occurred in a unique subset of maternal decidual cells previously known to express high levels of alkaline phosphatase and ADA. This induction occurred almost immediately after implantation in the periembryonic maternal decidual cells, shortly thereafter in antimesometrial decidual cells, and later in cells of the placental decidua basalis: all of which contain cell types thought to be undergoing programmed cell death. The expression of the pathway at the site of embryo implantation appears to be critical because its pharmacologic inhibition during pregnancy has been found to be embryolethal or teratogenic. Purine destruction at these nutritional interfaces (placenta and gastrointestinal tract) seem to override any potential economy of purine salvage, and may represent biochemical adaptation to nucleic acid breakdown occurring in the context of dietary digestion or extensive programmed cell death.

5'-Nucleotidase↗

PubMed2003

The first crystal structure of archaeal aldolase. Unique tetrameric structure of 2-deoxy-d-ribose-5-phosphate aldolase from the hyperthermophilic archaea Aeropyrum pernix.

A gene encoding a 2-deoxy-d-ribose-5-phosphate aldolase (DERA) homolog was identified in the hyperthermophilic Archaea Aeropyrum pernix. The gene was overexpressed in Escherichia coli, and the produced enzyme was purified and characterized. The enzyme is an extremely thermostable DERA; its activity was not lost after incubation at 100 degrees C for 10 min. The enzyme has a molecular mass of approximately 93 kDa and consists of four subunits with an identical molecular mass of 24 kDa. This is the first report of the presence of tetrameric DERA. The three-dimensional structure of the enzyme was determined by x-ray analysis. The subunit folds into an alpha/beta-barrel. The asymmetric unit consists of two homologous subunits, and a crystallographic 2-fold axis generates the functional tetramer. The main chain coordinate of the monomer of the A. pernix enzyme is quite similar to that of the E. coli enzyme. There was no significant difference in hydrophobic interactions and the number of ion pairs between the monomeric structures of the two enzymes. However, a significant difference in the quaternary structure was observed. The area of the subunit-subunit interface in the dimer of the A. pernix enzyme is much larger compared with the E. coli enzyme. In addition, the A. pernix enzyme is 10 amino acids longer than the E. coli enzyme in the N-terminal region and has an additional N-terminal helix. The N-terminal helix produces a unique dimer-dimer interface. This promotes the formation of a functional tetramer of the A. pernix enzyme and strengthens the hydrophobic intersubunit interactions. These structural features are considered to be responsible for the extremely high stability of the A. pernix enzyme. This is the first description of the structure of hyperthermophilic DERA and of aldolase from the Archaea domain.

Amino Acid Sequence↗