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PubMed1994

Crystal structure of a catalytic antibody Fab with esterase-like activity.

BACKGROUND: Antibodies with catalytic properties can be prepared by eliciting an antibody response against 'transition state analog' haptens. The specificity, rate and number of reaction cycles observed with these antibodies more closely resemble the properties of enzymes than any of the many other known enzyme-mimicking systems. RESULTS: We have determined to 3 A resolution the first X-ray structure of a catalytic antibody Fab. This antibody catalyzes the hydrolysis of a p-nitrophenyl ester. In conjunction with binding studies in solution, this structure of the uncomplexed site suggests a model for transition state fixation where two tyrosines mimic the oxyanion binding hole of serine proteases. A comparison with the structures of known Fabs specific for low molecular weight haptens reveals that this catalytic antibody has an unusually long groove at its combining site. CONCLUSION: Since transition state analogs contain elements of the desired product, product inhibition is a severe problem in antibody catalysis. The observation of a long groove at the combining site may relate to the ability of this catalytic antibody to achieve multiple cycles of reaction.

Antibodies, Monoclonal↗
PubMedDate not supplied

Nurse faculty practice: from theory to reality.

This article discusses the developmental stages involved in moving the concept of faculty practice from theory to reality. Bennis, Benne, and Chin's theories for planned change and Hage's theory for organizational change with a dominant coalition form the theoretical bases for this redical innovation. Smith's Time Line serves as the framework for tracing the development of the plan over its 6-year history. The political, logistic, and financial promoters and constraints at each stage of development are identified. Faculty reaction to the idea of faculty practice is presented, and the mechanisms used to bring about group consensus are described. The key elements for the successful transition of this plan are a small and resilient group of faculty planners and adaptation of existing organizational systems to accommodate nursing's interests. Finally, the practice plan is presented at its present stage of development with its future goals.

Attitude of Health Personnel↗
PubMed1994

Properties of isolated recombinant N and C domains of chicken troponin C.

The two globular N and C domains of chicken troponin C (TnC) are connected by an exposed alpha-helix (designated D/E; residues 86-94). Recombinant N (residues 1-90) and C (residues 88-162) domains containing either F29 or W29 and F105 or W105 have been engineered and expressed in Escherichia coli. These termination and initiation sites were chosen to minimize disruption of side-chain interactions between the D/E helix and other residues. W29 and W105 served as useful spectral probes for monitoring Ca(2+)-induced structural transitions of the N and C domains, respectively [Pearlstone et al. (1992) Biochemistry 31, 6545-6553; Trigo-Gonzalez et al. (1992) Biochemistry 31, 7009-7015]. By all criteria tested, the properties of the isolated F29W/N domain (1-90) were identical to those of the N domain in intact F29W. These included fluorescence emission spectra in the absence and presence of Ca2+/Mg2+, far-UV CD spectra, and Ca2+ affinity as monitored by fluorescence and ellipticity at 221 nm. Similar but not identical properties were observed for isolated F105W/C domain (88-162) and intact F105W. A summation of the far-UV CD spectra (+/- Ca2+) of the two domains was virtually superimposable on that of the intact protein. Of the total Ca(2+)-induced ellipticity change at 221 nm, 27% could be assigned to the N domain and 73% to the C domain. The data suggest a significant Ca(2+)-induced transition involving secondary structural elements of the N domain.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2001

Regular and inverse secondary kinetic enthalpy effects (KHE) for the rate of inversion of thioether and 1,1'-biisoquinoline complexes of ruthenium and osmium.

Thioether complexes with the formula Delta/Lambda-chloro(thioether)bis(2,2'-bipyridine)metal(II) (M = Ru, Os; thioether = dimethyl sulfide (3a(+)), diethyl sulfide (3b(+)), and tetrahydrothiophene (3c(+))) have been synthesized. The rates of inversion at the sulfur atom of the thioether ligands have been measured by spin-inversion transfer and line-shape NMR methods. In every case, the ruthenium derivative exhibits a faster inversion frequency at a given temperature than the corresponding osmium derivative. In contrast, similar complexes with the formula chloro(delta/lambda-1,1'-biisoquinoline)(2,2':6',2"-terpyridine)metal(II), 4(M=Ru,Os)(+), undergo atropisomerization of the misdirected 1,1'-biisoquinoline (1,1'-biiq) ligand with rates that are faster for osmium than ruthenium. As a result of the lanthanide contraction effect and the similar metric parameters associated with the structures of second-row and third-row transition metal derivatives, steric factors associated with the isomerizations are presumably similar for the Ru and Os derivatives of these compounds. Since third-row transition metal complexes tend to have larger bond dissociation enthalpies (BDE) than their second-row congeners, we conclude the difference in reactivities of 3(M=Ru)(+) versus 3(M=Os)(+) and 4(M=Ru)(+) versus 4(M=Os)(+) are attributed to electronic effects. For 3, the S3p lone pair of the thioether, the principal sigma donor orbital, is orthogonal to the metal sigma acceptor orbital in the transition state of inversion at sulfur and the S 3s orbital is an ineffective sigma donor. Thus, a regular relationship between the kinetic and thermodynamic stabilities of 3(M=Ru)(+) and 3(M=Os)(+) is observed for the directed <==> [misdirected] <==> directed (DMD) isomerization (the more thermodynamically stable bond is less reactive). In contrast, atropisomerization of 4(+) involves redirecting (strengthening) the M-N bonds of the misdirected 1,1'-biiq ligand in the transition state. Therefore, an inverse relationship between the kinetic and thermodynamic stabilities of 4(M=Ru)(+) and 4(M=Os)(+) is observed for the misdirected <==> [directed] <==> misdirected (MDM) isomerization (the more thermodynamically stable bond is more reactive). The rates obtained for 4(+) are consistent with the rates of atropisomerization of Delta/Lambda-(delta/lambda-1,1'-biisoquinoline)bis(2,2'-bipyridine)metal(II), 1(M=Ru,Os)(2+), and (eta(6)-benzene) Delta/Lambda-(delta/lambda-1,1'-biisoquinoline)halometal(II), 2(M=Ru,Os;halo=Cl,I)(+), that we reported previously. We term the relative rates of reaction of second-row versus third-row transition metal derivatives kinetic element effects (KEE = k(second)/k(third)). While the KEE appears to be generally useful when comparing reactions of isostructural species (e.g. the relative rates of 1(M=Ru)(2+), 1(M=Os)(2+), and 1(M=Ir)(3+)), different temperature dependencies of reactions prevent the comparison of related reactions between species that have different structures (e.g., the 1,1'-biiq atropisomerization reactions of 1(M=Ru,Os)(2+) versus 2(M=Ru,Os;halo=Cl,I)(+) versus 4(M=Ru,Os)(+)). This problem is overcome by comparing entropies of activation and kinetic enthalpy effects (KHE = DeltaH(third)/DeltaH(second)). For a given class of 1,1'-biiq complexes, we observe a structure/reactivity relationship between DeltaH and the torsional twist of the 1,1'-biiq ligands that are measured in the solid state.

Journal Article↗
PubMed2002

Cation substitutional chemistry in the r(7)x(12)z-type structure for four lanthanum iodide examples.

A series of quaternary lanthanide halide cluster compounds ALa(6)I(12)Z with transition metal interstitials (principally Os) and alkali or alkaline-earth metal cations (A = Na, Mg, Ca, Sr) have been synthesized by high temperature solid state techniques. The compounds were structurally characterized by single crystal and powder X-ray diffraction methods. The new compounds are isotypic (R3, Z = 3) with rhombohedral R(7)X(12)Z (R = Sc, Y, La-Lu; X = Cl, Br, I; Z = transition or main group element) and contain nominally octahedral R(6)X(12) units centered by interstitial Z. Here, the cation (A) rather than the seventh R occupies the isolated position between clusters along c as A(x)R(1-x) with 0 < x <or= 1. The balance of the structure remains essentially unchanged. The refined X-ray crystal structures of NaLa(6)I(12)Os, (Ca(0.906)La(0.094))La(6)I(12)Os, (Ca(0.621)La(0.379))La(6)I(12)Os, SrLa(6)I(12)Os, and (Sr(0.526)La(0.474))La(6)I(12)Os are reported, along with results of electrical resistivity and magnetic susceptibility measurements on these and some analogous phases. The phases are generally semiconductors, but the complex paramagnetic properties are not those of simple spin-only systems.

Journal Article↗
PubMed2006

Evolution of alternative transcriptional circuits with identical logic.

Evolution of gene regulation is an important contributor to the variety of life. Here, we analyse the evolution of a combinatorial transcriptional circuit composed of sequence-specific DNA-binding proteins that are conserved among all eukaryotes. This circuit regulates mating in the ascomycete yeast lineage. We first identify a group of mating genes that was transcriptionally regulated by an activator in a fungal ancestor, but is now transcriptionally regulated by a repressor in modern bakers' yeast. Despite this change in regulatory mechanism, the logical output of the overall circuit remains the same. By examining the regulation of mating in modern yeasts that are related to different extents, we deduce specific, sequential changes in both cis- and trans-regulatory elements that constitute the transition from positive to negative regulation. These changes indicate specific mechanisms by which fitness barriers were traversed during the transition.

Amino Acid Sequence↗
PubMed1998

Merger management: a challenge to nursing leadership.

AIM: The article reviews the merger process of two obstetric divisions. BACKGROUND: Mergers of acute care facilities are becoming common due to the need to move towards a market orientation. There is a growing emphasis on corporations, competition, and profit and cost control. Nursing leadership in managing merger processes is crucial. METHODS: Information originated from administrative decisions, feelings expressed by the staff, author observations, and statistical data. Data was analysed by illustrating the merger phases, compared to the literature and research studies. KEY ISSUES: Issues related to planning and preparing the process, the management of human resources, the development of organizational culture, and the physical changes, are vital. CONCLUSIONS: Planning ahead, involving all partners from the early stages, extensive dialogue among colleagues and strong nursing leadership are key elements for a smooth transition.

Decision Making, Organizational↗
PubMed2005

Multireference configuration interaction based electronic Floquet states for molecules in an intense radiation field: theory and application to Li(2)+.

A multireference configuration interaction (CI) method which includes single and double excitations based description of adiabatic Floquet states for the electronic structure of a molecule in an intense laser field is introduced. Using a variant of a recently introduced configuration state function (CSF) based Table-CI methodology, it is shown that the multiple states of several irreducible representations required for a good description of low-lying Floquet states can be obtained using modifications of computational molecular electronic structure techniques. In particular, formulas for all components of the transition dipole moment matrix elements within the CSF-based Table-CI method are derived and presented. Moreover, the flexibility of the recently introduced macroconfiguration description of model and external configuration spaces is shown to lead to multiple potential energy surfaces of sufficiently uniform quality to allow construction of useful Floquet states. The formalism and computer programs developed are demonstrated on Li(2) (+) in a 0.9 x 10(12) W/cm(2) field. In analogy with Na(2) (+), the 1,2 (2)Sigma(g) (+), 1,2 (2)Sigma(u) (+), 1 (2)Pi(g), and 1 (2)Pi(u) states are of relevance, although the pattern of couplings is shown to be more complex. A hitherto unnoticed metastable state, which correlates asymptotically with 2 (2)Sigma(u) (+), is described.

Journal Article↗
PubMed1996

Global properties of the mapping between local amino acid sequence and local structure in proteins.

Local protein structure prediction efforts have consistently failed to exceed approximately 70% accuracy. We characterize the degeneracy of the mapping from local sequence to local structure responsible for this failure by investigating the extent to which similar sequence segments found in different proteins adopt similar three-dimensional structures. Sequence segments 3-15 residues in length from 154 different protein families are partitioned into neighborhoods containing segments with similar sequences using cluster analysis. The consistency of the sequence-to-structure mapping is assessed by comparing the local structures adopted by sequence segments in the same neighborhood in proteins of known structure. In the 154 families, 45% and 28% of the positions occur in neighborhoods in which one and two local structures predominate, respectively. The sequence patterns that characterize the neighborhoods in the first class probably include virtually all of the short sequence motifs in proteins that consistently occur in a particular local structure. These patterns, many of which occur in transitions between secondary structural elements, are an interesting combination of previously studied and novel motifs. The identification of sequence patterns that consistently occur in one or a small number of local structures in proteins should contribute to the prediction of protein structure from sequence.

Amino Acid Sequence↗
PubMed1984

Generation and functional analyses for base-substitution mutants of the adenovirus 2 major late promoter.

The function of guanosine residues surrounding the TATA box of the adenovirus 2 major late promoter (MLP) in promoting efficient transcription initiation and in selecting a specific transcription start site in vitro has been examined. Multiple and single base substitutions (G----A) were generated in this region (from -63 to +25 relative to the cap site, +1) of the MLP. The promoter activities of the wild type and 21 mutants were assayed in an in vitro transcription system using whole cell extract (WCE) prepared from HeLa cells. The results suggest that the strings of G residues immediately adjacent to the TATA box are not required for full promoter activity in vitro. These G residues also appear not to be involved in the selection of a specific transcription start site by RNA polymerase II in vitro, since the identical cap site was used by wild-type and mutated MLP's. However, two G residues (-55 and -57) were identified as part of an upstream promoter element: a G----A transition at either -55 or -57 resulted in a 2.6 fold reduction in promoter activity. However, neither single nor double G----A transitions at -62 and -63 had an effect on promoter activity.

Adenoviruses, Human↗
PubMed1996

Gene transfer and therapeutic drug monitoring.

During the next decade, gene therapy will evolve from a medical curiosity into an essential component of medical practice. One of the elements necessary for this transition will be the development of simple and accurate ways of monitoring both the vectors used to transfer the genes of interest and the function of the genes themselves. This article reviews the difficulties in achieving these aims and describes ways in which the technology of gene transfer offers a novel means of monitoring current therapies.

Drug Monitoring↗
PubMed2000

Anisotropic vacuum-induced interference in decay channels

We demonstrate how the anisotropy of the vacuum of the electromagnetic field can lead to quantum interferences among the decay channels of close lying states. Our key result is that interferences are given by the scalar formed from the antinormally ordered electric field correlation tensor for the anisotropic vacuum and the dipole matrix elements for the two transitions. We present results for emission between two conducting plates as well as for a two photon process involving fluorescence produced under coherent cw excitation.

Journal Article↗
PubMed2005

Important role of blood rheology in atherosclerosis of patients with hemodialysis.

Concerning a role of blood rheology for atherosclerosis in patients with hemodialysis (HD), little data are available. It may be due to the fact that the method for evaluating rheologic properties of circulating blood has been limited. We examined blood rheology in 118 HD patients by using microchannel array flow analyzer that makes it possible to directly observe the flow of blood cell elements through the microchannel. Transit time (T(B)) of heparinized whole blood through slit pores (7 x 30 microm) was used as an index of rheology and related with various inflammatory biomarkers such as high-sensitive CRP (hsCRP), monocyte chemotactic protein-1, osteopontin, or fibrinogen (Fg). Moreover, as a surrogate marker of atherosclerosis, carotid intima-media thickness (IMT) and aortic stiffness evaluated by brachial-ankle pulse-wave velocity (baPWV) were studied. In HD patients, T(B) had strong positive correlations with hsCRP (r = 0.427; p < 0.00001), Fg (r = 0.452; p < 0.00001), and osteopontin (r = 0.227; p < 0.0134). Further, T(B) was significantly well correlated with IMT (r = 0.400; p < 0.0001) and PWV (r = 0.470; p < 0.0001). Multivariate regression analysis showed that baPWV, IMT, Fg, hematocrit, white blood cell count, and CRP were chosen as significant explanatory factors for T(B.) These results suggest that blood rheology may play an important role for atherosclerosis in patients with HD.

Adult↗
PubMed1998

Phenotypic, morphologic changes and Ig secretion induced on B-NHL cells in vitro by interferon alpha and all-trans-retinoic acid: possible progression toward a more differentiated state.

Twenty-five B-cell-nonHodgkin's lymphomas (B-NHL): 6 lymphocytic, 2 centrocytic, 13 follicular, centrocytic/centroblastic, 2 lymphoplasmocytoid and 2 centroblastic were tested for their ability to acquire features of mature plasma cells under the effect of interferon alpha (final concentration, 600 UI/ml), all-trans-retinoic-acid (ATRA) (final concentration, 10(-6) mol/l) and the association of both. B-NHL cells were negatively purified (>99%) by an immunomagnetic method, cultured for 7 d with or without interferon and ATRA, then stained with anti-CD19, CD20, surface Ig, DR, CD38 and with anti-CD138 (syndecan-1) antibody-recognizing plasma cells. Ig production was estimated in culture supernatants by an ELISA method and changes in cell morphology were investigated on May-Grunwald-Giemsa-stained cytospin preparations. In all cases interferon and ATRA, alone or in association, were able to induce changes in the immunophenotypic profile, associated or not with morphologic changes and induction of Ig secretion. All changes were greatly variable from one to the other B-NHL sample and no relationship could be found between a particular pattern of change and the histological subtype. Interferon alpha was more potent than ATRA in inducing changes. In favour of a differentiation process, we observed a concomitant decrease of DR expression and increase of CD38 expression in 8 cases with interferon alpha, and in 4 cases with ATRA. Although interferon- or ATRA-treated cells did not display cytologic, functional features and changes of the immunophenotypic profile fully compatible with those of terminally differentiated cells, these results suggest a possible transition toward more differentiated elements, especially with interferon alpha.

Antigens, Surface↗
PubMed1992

Inhibition of structural changes in the simian virus 40 core origin of replication by mutation of essential origin sequences.

Mutation of the simian virus 40 (SV40) origin of replication (ori) has revealed the presence of three critical domains needed for DNA replication. The outer two domains, the AT tract and early palindrome element (EP), colocalize with DNA regions that become structurally altered in the presence of the SV40 large tumor antigen (T antigen) and ATP. Mutations within each domain were examined for their effect on the distortion of ori DNA by T antigen, as assayed by the sensitivity of DNA to KMnO4 oxidation. We have found that mutations in the AT tract that inhibit SV40 DNA replication also inhibit the distortion of the AT tract. Similarly, mutations in the EP inhibited the generation of structural changes in this element by T antigen. Although AT-tract mutations or mutations on the late side of ori affected structural changes only in the AT tract, certain EP mutations or mutations on the early side of ori also inhibited AT-tract distortion. Mutation of the flanking regions did not significantly affect either the affinity of T antigen for ori or the rate of binding to ori. We conclude from these results that the primary function of the flanking ori domains is to undergo structural changes required during the initiation of SV40 DNA replication. Moreover, our results suggest that the efficiency of replication initiation is significantly affected by the degree to which the flanking elements undergo a structural transition.

Antigens, Polyomavirus Transforming↗
PubMed1994

Evidence for the ambiguity of the term constipation: the role of irritable bowel syndrome.

A satisfactory definition of constipation is elusive. An important and measurable element is slow colonic transit. Whole gut transit time, a proxy for colonic transit time, can be estimated from self recorded data on stool form and frequency. Our aim was to compare whole gut transit time with subjective definitions in the context of the general population. In a community based sample of 731 women aged 25-69 years the estimated whole gut transit time was compared with two subjective assessments of constipation-the woman's own perception and a symptom based definition proposed by an international working team (Rome definition). We have defined slow whole gut transit time as > 2 SD above the mean in women who seldom passed lumpy stools (that is, > 92 hours). Slow transit was present in 9.3% of the sample. Similar numbers met the subjective definitions (8.5% and 8.2%). However, the overlap between the three definitions was poor. Of 68 women with estimated slow transit, 28 had self perceived constipation, 20 had Rome defined constipation, and only 11 had both. Of subjects classified as constipated by the subjective definitions only 37% had slow transit; they had a high prevalence of irritable bowel symptoms. In conclusion, this study showed that the term constipation is ambiguous and often misleading and that attempts to base a definition on symptoms are misguided. In epidemiological studies, conclusions about the prevalence of constipation should be based on records of stool type and timing.

Adult↗
PubMed1995

CD34+ human hemopoietic progenitor cells of the bone marrow differ from those of the peripheral blood: an immunocytochemical and morphometric study.

CD34+ progenitor cells were harvested from bone marrow and peripheral blood from 10 healthy donors by immunomagnetic isolation and enrichment procedures. The CD34+ cell population was investigated using a battery of enzyme reactions and monoclonal antibodies on cytospin preparations. Additionally, morphometric measurements were carried out and also liquid suspension culture studies were performed to ascertain vitality and stem cell character. More than 95% of the total yield of medullary CD34 progenitors expressed CD45 (LCA), CD43 (MT1) and beta-glucuronidase. Reactivity with CD33 (My9), CD15 (LeuM1), CD38 (Leu17), CD20 (L26) and Ret40f (glycophorin C) was assumed to be in keeping with a transition into more differentiated elements of the various hemopoietic lineages. Morphometric analysis revealed conspicuous heterogeneity of the CD34+ cell population considering size measurements. This finding was in line with the diversities of antigen expression, indicating the more committed nature of CD34+ stem cells derived from the bone marrow in comparison with those progenitors isolated from the peripheral blood. Moreover, proliferation marker staining by PCNA disclosed a positivity in a considerable number of progenitors in contrast to the findings in CD34+ cells that are found in the peripheral blood.

Antibodies, Monoclonal↗