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1.
Heart Lung Circ ; 33(4): 420-442, 2024 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-38570258

RESUMEN

Over the past 5 years, early diagnosis of and new treatments for cardiac amyloidosis (CA) have emerged that hold promise for early intervention. These include non-invasive diagnostic tests and disease modifying therapies. Recently, CA has been one of the first types of cardiomyopathy to be treated with gene editing techniques. Although these therapies are not yet widely available to patients in Australia and New Zealand, this may change in the near future. Given the rapid pace with which this field is evolving, it is important to view these advances within the Australian and New Zealand context. This Consensus Statement aims to update the Australian and New Zealand general physician and cardiologist with regards to the diagnosis, investigations, and management of CA.


Asunto(s)
Amiloidosis , Cardiomiopatías , Consenso , Humanos , Amiloidosis/terapia , Amiloidosis/diagnóstico , Australia , Cardiomiopatías/terapia , Cardiomiopatías/diagnóstico , Nueva Zelanda
2.
Heart Lung Circ ; 29(7): 1054-1062, 2020 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-31594721

RESUMEN

BACKGROUND: Acute decompensated heart failure (ADHF) carries a high event rate following discharge. The complex interplay between age, frailty and decongestion may lend itself to a functional test. METHODS: In the doorbell test the patient simulates answering the doorbell. They are timed rising from a recumbent position, bending over twice and walking 10 metres, this time is added to the change in respiratory rate. We aimed to determine if the doorbell test was associated with post ADHF events (death or readmission). The test was performed at hospital discharge, with follow up at 30-days and 1-year. RESULTS: In 74 patients at 30-days there was a 14% event rate. At 1-year there were 40 (54%) events (9 deaths and 31 readmissions, 28 were cardiovascular of which 14 were [heart failure] HF). Amongst those who had an event at 30-days only doorbell test scores were different (58 [36,72] vs 32 [26,53] p < 0.05). One-year (1-year) events were associated with doorbell test scores (47 [29,62] vs 30 [26,42] p < 0.05), body weight (78 kg [68,94] vs 95 [76,105] (p < 0.05), creatinine (134 mmol/L [114, 173] vs 99 [82, 133] p < 0.01) and age (76 years [61,86] vs 67 [53, 73] p < 0.01). Heart failure readmissions were associated with doorbell test scores (56 [46,68] vs 30 [26,47] p < 0.001). Death was associated with body weight (74 kg [69,81] vs 88 [72,101] p < 0.05) and age (83 years [78,86] vs 69 [55,77] p < 0.01). After age stratification, the hazard ratio for heart failure readmission associated with a high doorbell test score was 11.08 (95%C.I. 2.01-61.17 p = 0.006), while the hazard ratio for 1-year cardiovascular readmission was 4.62 (95%C.I. 1.71-12.51 p = 0.003). There was no association with 1-year mortality. CONCLUSION: The doorbell test represents a novel test of multiple domains of the ADHF pre-discharge state and demonstrates an association with 30-day and 1-year rehospitalisation.


Asunto(s)
Prueba de Esfuerzo/métodos , Fragilidad/diagnóstico , Insuficiencia Cardíaca/diagnóstico , Caminata/fisiología , Enfermedad Aguda , Anciano , Femenino , Fragilidad/complicaciones , Fragilidad/fisiopatología , Insuficiencia Cardíaca/complicaciones , Insuficiencia Cardíaca/fisiopatología , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Factores de Riesgo
3.
J Chem Phys ; 142(24): 244705, 2015 Jun 28.
Artículo en Inglés | MEDLINE | ID: mdl-26133447

RESUMEN

We employ Monte Carlo simulation to examine encapsulation in a system comprising Janus oblate spheroids and isotropic spheres. More specifically, the impact of variations in temperature, particle size, inter-particle interaction range, and strength is examined for a system in which the spheroids act as the encapsulating agents and the spheres as the encapsulated guests. In this picture, particle interactions are described by a quasi-square-well patch model. This study highlights the environmental adaptation and selectivity of the encapsulation system to changes in temperature and guest particle size, respectively. Moreover, we identify an important range in parameter space where encapsulation is favored, as summarized by an encapsulation map. Finally, we discuss the generalization of our results to systems having a wide range of particle geometries.

4.
Diabetes Care ; 47(1): 140-143, 2024 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-37988720

RESUMEN

OBJECTIVE: Ketoacidosis induced by sodium-glucose cotransporter 2 inhibitor (SGLT2i) treatment has been consistently observed in clinical practice in patients with type 2 diabetes despite minimal indication from the landmark cardiovascular outcome trials. It has been postulated that individuals without diabetes will not develop this complication due to an adequate insulin secretory capacity, which will protect against significant ketone formation. Cardiovascular outcome trials examining SGLT2i use in individuals with heart failure but not diabetes have not reported ketoacidosis. RESEARCH DESIGN AND METHODS: We describe the first two case reports of severe nondiabetic ketoacidosis after initiation of an SGLT2i for the treatment of heart failure with reduced ejection fraction, and we describe the management strategies employed and implication for the pathophysiology of SGLT2i-associated ketoacidosis. RESULTS: Each individual presented with ketoacidosis triggered by reduced oral nutrition intake. For both individuals, ketoacidosis resolved with intravenous glucose administration, encouragement of consumption of oral glucose-containing fluid, and minimal insulin administration. CONCLUSIONS: These two cases demonstrate that SGLT2i-associated ketoacidosis is possible in individuals without diabetes.


Asunto(s)
Diabetes Mellitus Tipo 2 , Cetoacidosis Diabética , Insuficiencia Cardíaca , Cetosis , Inhibidores del Cotransportador de Sodio-Glucosa 2 , Humanos , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hipoglucemiantes/efectos adversos , Cetoacidosis Diabética/inducido químicamente , Inhibidores del Cotransportador de Sodio-Glucosa 2/efectos adversos , Volumen Sistólico , Insulina/efectos adversos , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/complicaciones , Glucosa/uso terapéutico , Sodio/efectos adversos
5.
Langmuir ; 29(27): 8517-23, 2013 Jul 09.
Artículo en Inglés | MEDLINE | ID: mdl-23742624

RESUMEN

In self-assembly, the anisotropy of the building blocks and their formation of complex structures have been the subject of considerable recent research. Extending recent research on Janus particles and completing the study of Janus spheroids, we conduct a systematic investigation on the self-assembly of Janus prolate spheroids based on a primitive model that we proposed. Janus prolate spheroids are particles that have a prolate spheroidal body and two hemi-surfaces along the major axis coded with different chemical properties. Using Monte Carlo simulations, we investigate the effects of the aspect ratio on the self-assembly process. In contrast to the vesicle-like aggregates for Janus oblate spheroids, we obtain various ordered cluster structures for Janus prolate spheroids through self-assembly. With an increasing aspect ratio, we find a transition of cluster morphology, from vesicles to tubular micelles and micelles. In particular, a relatively small change in the aspect ratio leads to a rather significant change in morphology. We apply a cluster analysis to understand the mechanism associated with such a transition.

6.
Langmuir ; 29(14): 4470-6, 2013 Apr 09.
Artículo en Inglés | MEDLINE | ID: mdl-23484544

RESUMEN

Extending recent work on a generic bipolar model proposed to study the nanochain formation of amelogenin molecules, we conduct a systematic investigation in this paper on the self-assembly of such a model via sweeping the relative parameter space. The bipolar model consists of a short-range attraction and an off-center Coulomb repulsion for the supermolecule. Through the Brownian dynamics simulation of both translational and rotational motions, we study the kinetics of the self-assembly and the structure of clusters formed within the system for various interaction settings. From the results of structure factor and cluster analysis, we find that the range of the repulsive interaction has a sensitive impact in controlling the cluster size, while the strength of the attractive interaction dominates the cluster morphology such that the greater the attraction among particles, the more elongated the cluster formed.


Asunto(s)
Amelogenina/química , Modelos Moleculares , Amelogenina/metabolismo , Interacciones Hidrofóbicas e Hidrofílicas , Movimiento , Conformación Proteica
7.
Langmuir ; 28(1): 3-9, 2012 Jan 10.
Artículo en Inglés | MEDLINE | ID: mdl-22171980

RESUMEN

We propose a primitive model of Janus ellipsoids that represents particles with an ellipsoidal core and two semisurfaces coded with dissimilar properties, for example, hydrophobicity and hydrophilicity, respectively. We investigate the effects of the aspect ratio on the self-assembly morphology and aggregation processes using Monte Carlo simulations. We also discuss certain differences between our results and those of earlier results for Janus spheres. In particular, we find that the size and structure of the aggregate can be controlled by the aspect ratio.

8.
J Phys Chem B ; 112(26): 7803-9, 2008 Jul 03.
Artículo en Inglés | MEDLINE | ID: mdl-18537289

RESUMEN

Anisotropic protein interactions play a role in globular protein phase transitions, including the shape of fluid-fluid coexistence curves and the formation of hemoglobin polymer fibers in sickle cell disease. Also, the solvent has been shown to play an important role in the phase behavior of some aqueous protein solutions, through the release or trapping of water molecules upon crystallization. Both anisotropy and solvent effects have been treated separately in earlier theoretical studies. Here we propose and analyze a simple, composite model that treats both anisotropy and solvent effects. We find that this model qualitatively explains some phase behavior that has been observed in protein solutions, including normal and retrograde solubility curves and lower critical points.


Asunto(s)
Pliegue de Proteína , Proteínas/química , Proteínas/metabolismo , Solventes/química , Anisotropía , Modelos Biológicos
9.
Int J Cardiol ; 254: 230-237, 2018 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-29407096

RESUMEN

BACKGROUND: The central role of left ventricular ejection fraction (LVEF) as the definitive risk marker of adverse outcomes in ischemic and nonischemic cardiomyopathy is increasingly uncertain. The current study aimed to conduct a systematic review and meta-analysis with the objective of evaluating the prognostic importance of Late Gadolinium Enhancement (LGE) in ischemic cardiomyopathy (ICM) and non-ischemic cardiomyopathy (NICM) on the key endpoints of all-cause mortality, cardiovascular mortality and sudden death. METHODS: The study was prospectively registered in PROPSERO (CRD 42016039034). Electronic databases and reference lists were searched for studies evaluating the impact of LGE-CMR on all-cause mortality, cardiovascular mortality, ventricular arrhythmia or sudden death, or major adverse cardiovascular events. Data were extracted from 36 studies including n=7882 patients. RESULTS: LGE was strongly associated with all-cause mortality HR 2.96 (95%CI: 2.37, 3.70, P<0.001), cardiovascular mortality HR 3.27 (95% CI: 2.05, 5.22, P<0.001), ventricular arrhythmia and sudden cardiac death HR 3.76 (95% CI: 3.14, 4.52, P<0.001), and major adverse cardiovascular events HR 3.24 (95% CI: 2.32, 4.52, P<0.001). In subgroup analyses, LGE was associated with all-cause mortality and cardiovascular mortality in both LVEF≤35% and LVEF>35% patients (P<0.001 all endpoints), as well as in nonischemic and ischemic cardiomyopathy. CONCLUSION: Late Gadolinium Enhancement (LGE) in CMR predicts all-cause mortality, cardiovascular mortality, ventricular arrhythmia and sudden death, and major adverse cardiovascular events, independent of LVEF. Future trials of investigational therapies in NICM and ICM should consider the utilization of LGE to identify patients at risk of adverse outcomes.


Asunto(s)
Cardiomiopatías/diagnóstico por imagen , Cardiomiopatías/mortalidad , Muerte Súbita Cardíaca/epidemiología , Gadolinio , Isquemia Miocárdica/diagnóstico por imagen , Isquemia Miocárdica/mortalidad , Estudios de Cohortes , Humanos , Mortalidad/tendencias , Ensayos Clínicos Controlados Aleatorios como Asunto/métodos
10.
J Phys Chem B ; 111(6): 1478-81, 2007 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-17243665

RESUMEN

The liquid-liquid phase separation curves for lysozyme in a salt solution are known to depend on salt type and salt concentration. For the case of monovalent cations, the cloud point temperature typically increases with increasing salt concentration, for fixed lysozyme concentration. For the case of divalent cations, however, a maximum in the cloud point temperature is observed that has been interpreted as being due to ion binding to the protein surface and subsequent water structuring. In this paper, we use a simple square well model due to Grigsby et al. (Biophys. Chem. 2001, 91, 231-243), whose well depth depends on salt type and salt concentration, to determine the phase coexistence surfaces from experimental data. The surfaces are shown as a function of temperature, salt concentration, and protein concentration for two typical salts, NaCl and MgCl2. These surfaces are calculated using the results of a single standard Monte Carlo simulation and a simple scaling argument and are in reasonably good agreement with known experimental results.

11.
J Clin Med ; 5(8)2016 Aug 17.
Artículo en Inglés | MEDLINE | ID: mdl-27548237

RESUMEN

Current guidelines recommend initiation of a P2Y12 inhibitor for all patients with non-ST-elevation acute coronary syndrome (NSTE-ACS) at the time of diagnosis (pre-treatment); however, there are no randomized trials directly comparing pre-treatment with initiation at the time of angiography to support this practice. We explore clinical and institutional parameters potentially associated with benefit with this strategy in a decision-analytic model based on available evidence from randomised trials. A decision analysis model was constructed comparing three P2Y12 inhibitors in addition to aspirin in patients with NSTE-ACS. Based on clinical trial data, the cumulative probability of 30 day mortality, myocardial infarction (MI) and major bleeding were determined, and used to calculate the net clinical benefit (NCB) with and without pre-treatment. Sensitivity analysis was performed to assess the relationship between NCB and baseline ischemic risk, bleeding risk, time to angiography and local surgical revascularization rates. Pre-treatment with ticagrelor and clopidogrel was associated with a greater than 50% likelihood of providing a >1% increase in 30 day NCB when baseline estimated ischemic risk exceeds 11% and 14%, respectively. Prasugrel pre-treatment did not achieve a greater than 50% probability of an increase in NCB regardless of baseline ischemic risk. Institutional surgical revascularization rates and time to coronary angiography did not correlate with the likelihood of benefit from P2Y12 pre-treatment. In conclusion, pre-treatment with P2Y12 inhibition is unlikely to be beneficial to the majority of patients presenting with NSTE-ACS. A tailored assessment of each patient's individual ischemic and bleeding risk may identify those likely to benefit.

12.
Philos Trans A Math Phys Eng Sci ; 368(1933): 5619-32, 2010 Dec 28.
Artículo en Inglés | MEDLINE | ID: mdl-21078638

RESUMEN

We study the effect that the heterogeneity present among the elements of an ensemble of coupled excitable neurons has on the collective response of the system to an external signal. We consider two different interaction scenarios, one in which the neurons are diffusively coupled and another in which the neurons interact via pulse-like signals. We find that the type of interaction between the neurons has a crucial role in determining the response of the system to the external modulation. We develop a mean-field theory based on an order parameter expansion that quantitatively reproduces the numerical results in the case of diffusive coupling.


Asunto(s)
Modelos Neurológicos , Neuronas/fisiología , Algoritmos , Animales , Biodiversidad , Biofisica/métodos , Simulación por Computador , Difusión , Humanos , Modelos Biológicos , Modelos Estadísticos , Modelos Teóricos , Oscilometría
13.
Plasmid ; 59(2): 86-101, 2008 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-18192011

RESUMEN

Conjugative plasmids have evolved entry exclusion mechanisms to inhibit redundant DNA transfer from donor cells into recipients harboring isogenic or closely related plasmids. This exclusion phenomenon has been documented in the incompatibility H group (IncH) plasmid R27. A cosmid library representing the majority of the large (180kb) R27 plasmid was transformed into recipient cells and a conjugation assay identified that an operon located in the conjugative transfer region 2 (Tra2) of R27, the Z operon, mediated entry exclusion in the IncH plasmid. Reverse-transcriptase analysis revealed that the Z operon is comprised of four genes, 015, eexB, 017, and eexA. Sub-cloning of the individual genes located within the Z operon and subsequent screening for the entry exclusion phenotype determined that two genes, eexA and eexB, independently inhibit the entry of IncH-related plasmids. Bacterial fractionation studies predominantly localized the EexA protein to the cytoplasmic membrane, and the EexB protein to the outer membrane. Recipient cells expressing EexA and EexB were unable to exclude the entry of R27 plasmids harboring mutations within the IncH entry exclusion genes eexA and eexB. The IncH entry exclusion proteins EexA and EexB likely prevent redundant plasmid transfer by interaction with one another.


Asunto(s)
Conjugación Genética/fisiología , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Proteínas de la Membrana/metabolismo , Plásmidos/metabolismo , Membrana Celular/metabolismo , Conjugación Genética/genética , Cósmidos , Escherichia coli/citología , Escherichia coli/genética , Escherichia coli/ultraestructura , Sistemas de Lectura Abierta/genética , Operón/genética , Transporte de Proteínas , ADN Polimerasa Dirigida por ARN/metabolismo , Temperatura , Transcripción Genética
14.
Microbiology (Reading) ; 153(Pt 2): 428-441, 2007 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-17259614

RESUMEN

Bacterial conjugation is a DNA transfer event that requires three plasmid-encoded multi-protein complexes: the membrane-spanning mating pair formation (Mpf) complex, the cytoplasmic nucleoprotein relaxosome complex, and a homo-multimeric coupling protein that links the Mpf and relaxosome at the cytoplasmic membrane. Bacterial two-hybrid (BTH) technology and immunoprecipitation were used to demonstrate an interaction between the IncH plasmid-encoded transfer protein TraJ and the coupling protein TraG. TraJ is essential for conjugative transfer but is not required for the formation of the conjugative pilus, and is therefore not regarded as an Mpf component. Fractionation studies indicated that TraJ shared a similar cellular domain to that of TraG at the cellular membrane. Protein blast analyses have previously identified TraJ homologues encoded in a multitude of plasmid and chromosomal genomes that were also found to encode an adjacent TraG homologue, thus indicating co-inheritance. BTH analysis of these TraJ and cognate TraG homologues demonstrated conservation of the TraJ-TraG interaction. Additional occurrences of the traJ-traG module were also detected in genomic sequence data throughout the Proteobacteria, and phylogenetic comparison of these IncH-like TraG proteins with the coupling proteins encoded by other conjugative transfer systems (including IncP, IncW and IncF) that lack TraJ homologues indicated that the H-like coupling proteins were distinct. Accordingly, the IncP, IncW and IncF coupling proteins were unable to interact with TraJ, but were able to interact with IncH plasmid-encoded TrhB, an Mpf component known to complex with its cognate coupling protein TraG. The divergence of the IncH-type coupling proteins may partly be due to the requirement of TraJ interaction, and notably, TraG and TraJ cumulatively represent the domain architecture of the known translocase family FtsK/SpoIIIE. It is proposed that TraJ is a functional part of the IncH-type coupling protein complex required for translocation of DNA through the cytoplasmic membrane.


Asunto(s)
Proteínas de la Membrana Bacteriana Externa/metabolismo , Membrana Celular/metabolismo , Conjugación Genética , ADN Bacteriano/metabolismo , Proteínas de Escherichia coli/metabolismo , Proteínas de la Membrana/metabolismo , Plásmidos/genética , Proteínas de la Membrana Bacteriana Externa/genética , Transporte Biológico , Cromosomas Bacterianos/enzimología , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Inmunoprecipitación , Proteínas de la Membrana/genética , Técnicas del Sistema de Dos Híbridos
15.
Phys Rev Lett ; 97(19): 194101, 2006 Nov 10.
Artículo en Inglés | MEDLINE | ID: mdl-17155633

RESUMEN

We present conclusive evidence showing that different sources of diversity, such as those represented by quenched disorder or noise, can induce a resonant collective behavior in an ensemble of coupled bistable or excitable systems. Our analytical and numerical results show that when such systems are subjected to an external subthreshold signal, their response is optimized for an intermediate value of the diversity. These findings show that intrinsic diversity might have a constructive role and suggest that natural systems might profit from their diversity in order to optimize the response to an external stimulus.

16.
Microbiology (Reading) ; 151(Pt 11): 3549-3561, 2005 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-16272378

RESUMEN

Bacterial conjugation is a horizontal gene transfer event mediated by the type IV secretion system (T4SS) encoded by bacterial plasmids. Within the T4SS, the coupling protein plays an essential role in linking the membrane-associated pore-forming proteins to the cytoplasmic, DNA-processing proteins. TraG is the coupling protein encoded by the incompatibility group HI plasmids. A hallmark feature of the IncHI plasmids is optimal conjugative transfer at 30 degrees C and an inability to transfer at 37 degrees C. Transcriptional analysis of the transfer region 1 (Tra1) of R27 has revealed that traG is transcribed in a temperature-dependent manner, with significantly reduced levels of expression at 37 degrees C as compared to expression at 30 degrees C. The R27 coupling protein contains nucleoside triphosphate (NTP)-binding domains, the Walker A and Walker B boxes, which are well conserved among this family of proteins. Site-specific mutagenesis within these motifs abrogated the conjugative transfer of R27 into recipient cells. Mutational analysis of the TraG periplasmic-spanning residues, in conjunction with bacterial two-hybrid and immunoprecipitation analysis, determined that this region is essential for a successful interaction with the T4SS protein TrhB. Further characterization of TraG by immunofluorescence studies revealed that the R27 coupling protein forms membrane-associated fluorescent foci independent of R27 conjugative proteins. These foci were found at discrete positions within the cell periphery. These results allow the definition of domains within TraG that are involved in conjugative transfer, and determination of the cellular location of the R27 coupling protein.


Asunto(s)
Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Factores R/genética , Fracciones Subcelulares/metabolismo , Secuencia de Aminoácidos , Conjugación Genética , Escherichia coli/genética , Escherichia coli/crecimiento & desarrollo , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Inmunoprecipitación , Proteínas de la Membrana/genética , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Temperatura , Transcripción Genética , Técnicas del Sistema de Dos Híbridos
17.
J Chem Phys ; 122(23): 234911, 2005 Jun 15.
Artículo en Inglés | MEDLINE | ID: mdl-16008491

RESUMEN

The properties of the solvent affect the behavior of the solution. We propose a model that accounts for the contribution of the solvent free energy to the free energy of globular proteins in solution. For the case of an attractive square-well potential, we obtain an exact mapping of the phase diagram of this model without solvent to the model that includes the solute-solvent contribution. In particular we find for appropriate choices of parameters upper critical points, lower critical points, and even closed loops with both upper and lower critical points similar to those found before [Macromolecules 36, 5843 (2003)]. In the general case of systems whose interactions are not attractive square wells, this mapping procedure can be a first approximation to understand the phase diagram in the presence of solvent. We also present simulation results for both the square-well model and a modified Lennard-Jones model.


Asunto(s)
Modelos Químicos , Modelos Moleculares , Proteínas/química , Proteínas/ultraestructura , Solventes/química , Simulación por Computador , Transición de Fase , Conformación Proteica , Soluciones
18.
J Chem Phys ; 120(17): 8318-26, 2004 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-15267753

RESUMEN

A continuum model of globular proteins proposed by Talanquer and Oxtoby [J. Chem. Phys. 109, 223 (1998)] is investigated numerically, with particular emphasis on the region near the metastable fluid-fluid coexistence curve. Classical nucleation theory is shown to be invalid not only in the vicinity of the metastable critical point but also close to the liquidus line. An approximate analytic solution is also presented for the shape and properties of the nucleating crystal droplet.


Asunto(s)
Cristalografía por Rayos X/métodos , Proteínas/química , Fenómenos Biofísicos , Biofisica , Fenómenos Químicos , Química Física , Simulación por Computador , Cristalización , Modelos Biológicos , Modelos Estadísticos , Termodinámica
19.
Mol Microbiol ; 49(1): 105-16, 2003 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-12823814

RESUMEN

Assemblies of plasmid-encoded proteins direct the conjugative transfer of plasmid DNA molecules between bacteria. These include the membrane-associated mating pair formation (Mpf) complex necessary for pilus production and the cytoplasmic relaxosome required for DNA processing. The proposed link between these distinct protein complexes is the coupling protein (the TraG family of proteins). Interactions between the coupling protein and relaxosome components have been previously characterized and we document here, for the first time, a direct interaction between the coupling protein and an Mpf protein. Using the adenylate cyclase bacterial two-hybrid (BTH) system, we present in vivo evidence that the IncHI1 plasmid R27-encoded proteins TraG and TrhB interact. This interaction was verified through a co-immunoprecipitation reaction. We have also been able to delineate the interaction domain of TrhB to TraG by showing a positive interaction using the first 220 amino acids of TrhB (452 aa). TrhB has a proline-rich domain from amino acids 135-173 which may serve to facilitate protein interactions and/or periplasmic extension. TrhB self association was detected using far-Western, co-immunoprecipitation, and also BTH analysis, which was used to define the homotypic interaction domain, comprising a predicted coiled-coil region at residues 77-124 of TrhB. These data support a model in which the coupling protein interacts with an Mpf component to target the transferring DNA strand held by the relaxosome to the transmembrane Mpf complex.


Asunto(s)
Proteínas Portadoras/metabolismo , Proteínas de Escherichia coli/metabolismo , Proteínas de la Membrana/metabolismo , Plásmidos/metabolismo , Transporte Biológico/fisiología , Proteínas Portadoras/genética , Conjugación Genética , Escherichia coli/genética , Escherichia coli/fisiología , Proteínas de Escherichia coli/genética , Sustancias Macromoleculares , Proteínas de la Membrana/genética , Plásmidos/genética , Conformación Proteica , Proteínas Recombinantes de Fusión/metabolismo , Técnicas del Sistema de Dos Híbridos
20.
J Bacteriol ; 184(8): 2173-80, 2002 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-11914349

RESUMEN

The conjugative transfer region 1 (Tra1) of the IncHI1 plasmid R27 was subjected to DNA sequence analysis, mutagenesis, genetic complementation, and an H-pilus-specific phage assay. Analysis of the nucleotide sequence indicated that the Tra1 region contains genes coding for mating pair formation (Mpf) and DNA transfer replication (Dtr) and a coupling protein. Insertional disruptions of 9 of the 14 open reading frames (ORFs) in the Tra1 region resulted in a transfer-deficient phenotype. Conjugative transfer was restored for each transfer mutant by genetic complementation. An intergenic region between traH and trhR was cloned and mobilized by R27, indicating the presence of an origin of transfer (oriT). The five ORFs immediately downstream of the oriT region are involved in H-pilus production, as determined by an H-pilus-specific phage assay. Three of these ORFs encode proteins homologous to Mpf proteins from IncF plasmids. Upstream of the oriT region are four ORFs required for plasmid transfer but not H-pilus production. TraI contains sequence motifs that are characteristic of relaxases from the IncP lineage but share no overall homology to known relaxases. TraJ contains both an Arc repressor motif and a leucine zipper motif. A putative coupling protein, TraG, shares a low level of homology to the TraG family of coupling proteins and contains motifs that are important for DNA transfer. This analysis indicates that the Mpf components of R27 share a common lineage with those of the IncF transfer system, whereas the relaxase of R27 is ancestrally related to that of the IncP transfer system.


Asunto(s)
Conjugación Genética , Plásmidos , Secuencia de Bases , Clonación Molecular , Prueba de Complementación Genética , Datos de Secuencia Molecular , Análisis de Secuencia de ADN
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