scholarly journals Oligomerization Profile of Human Transthyretin Variants with Distinct Amyloidogenicity

Molecules ◽  
2020 ◽  
Vol 25 (23) ◽  
pp. 5698
Author(s):  
Ana Frangolho ◽  
Bruno E. Correia ◽  
Daniela C. Vaz ◽  
Zaida L. Almeida ◽  
Rui M. M. Brito
Keyword(s):  
Early Stage ◽  
Clinical Manifestations ◽  
Attr Amyloidosis ◽  
Amyloid Fibril Formation ◽  
Acidic Conditions ◽  
Novel Therapeutic Strategies ◽  
Amyloid Diseases ◽  
Kinetics Of ◽  
Identification And Characterization

One of the molecular hallmarks of amyloidoses is ordered protein aggregation involving the initial formation of soluble protein oligomers that eventually grow into insoluble fibrils. The identification and characterization of molecular species critical for amyloid fibril formation and disease development have been the focus of intense analysis in the literature. Here, using photo-induced cross-linking of unmodified proteins (PICUP), we studied the early stages of oligomerization of human transthyretin (TTR), a plasma protein involved in amyloid diseases (ATTR amyloidosis) with multiple clinical manifestations. Upon comparison, the oligomerization processes of wild-type TTR (TTRwt) and several TTR variants (TTRV30M, TTRL55P, and TTRT119M) clearly show distinct oligomerization kinetics for the amyloidogenic variants but a similar oligomerization mechanism. The oligomerization kinetics of the TTR amyloidogenic variants under analysis showed a good correlation with their amyloidogenic potential, with the most amyloidogenic variants aggregating faster (TTRL55P > TTRV30M > TTRwt). Moreover, the early stage oligomerization mechanism for these variants involves stepwise addition of monomeric units to the growing oligomer. A completely different behavior was observed for the nonamyloidogenic TTRT119M variant, which does not form oligomers in the same acidic conditions and even for longer incubation times. Thorough characterization of the initial steps of TTR oligomerization is critical for better understanding the origin of ATTR cytotoxicity and developing novel therapeutic strategies for the treatment of ATTR amyloidosis.

A transitional stage in the commitment of mesoderm to hematopoiesis requiring the transcription factor SCL/tal-1

Development ◽  
2000 ◽  
Vol 127 (11) ◽  
pp. 2447-2459 ◽  
Author(s):  
S.M. Robertson ◽  
M. Kennedy ◽  
J.M. Shannon ◽  
G. Keller
Keyword(s):  
Transcription Factor ◽  
Embryoid Body ◽  
Early Stage ◽  
Es Cells ◽  
Blast Cell ◽  
Stage Of Development ◽  
Erythroid Precursors ◽  
Identification And Characterization ◽  
Definitive Hematopoiesis

In this report, we describe the identification and characterization of an early embryoid body-derived colony, termed the transitional colony, which contains cell populations undergoing the commitment of mesoderm to the hematopoietic and endothelial lineages. Analysis of individual transitional colonies indicated that they express Brachyury as well as flk-1, SCL/tal-1, GATA-1, (beta)H1 and (beta)major reflecting the combination of mesodermal, hematopoietic and endothelial populations. This pattern differs from that found in the previously described hemangioblast-derived blast cell colonies in that they typically lacked Brachyury expression, consistent with their post-mesodermal stage of development (Kennedy, M., Firpo, M., Choi, K., Wall, C., Robertson, S., Kabrun, N. and Keller, G. (1997) Nature 386, 488–493). Replating studies demonstrated that transitional colonies contain low numbers of primitive erythroid precursors as well as a subset of precursors associated with early stage definitive hematopoiesis. Blast cell colonies contain higher numbers and a broader spectrum of definitive precursors than found in the transitional colonies. ES cells homozygous null for the SCL/tal-1 gene, a transcription factor known to be essential for development of the primitive and definitive hematopoietic systems, were not able to form blast colonies but did form transitional colonies. Together these findings suggest that the transitional colony represents a stage of development earlier than the blast cell colony and one that uniquely defines the requirement for a functional SCL/tal-1 gene for the progression to hematopoietic commitment.

Characterization of Type B Carbonate Apatite Sintered Bodies

2011 ◽  
Vol 1301 ◽  
Author(s):  
N. Watanobe ◽  
T. Yoshioka ◽  
T. Ikoma ◽  
T. Kuwayama ◽  
T. Higaki ◽  
...  
Keyword(s):  
Dissolution Kinetics ◽  
Carbonate Apatite ◽  
Artificial Bone ◽  
Type B ◽  
Desorption Process ◽  
Different Temperatures ◽  
Acidic Conditions ◽  
Kinetics Of ◽  
Potential Use

ABSTRACTThe dissolution kinetics of type B carbonate apatite (CAp) disks sintered at different temperatures were investigated under acidic conditions similar to an osteoclastic desorption process in bone remodeling. The type B CAp disks, which were uniaxially pressed at 98MPa and sintered at temperatures of 600 to 700 ºC, showed single crystalline phase and a high relative density of 60-62 % compared to the stoichiometric density of 3.16g/cm3 of hydroxyapatite (HAp). The dissolution rate of type B CAp disks sintered at 650 ºC was 3.5 times faster than that of HAp disks at 650 ºC. These results indicate that the Type B CAp sintered disks show potential use as a biodegradable material for artificial bone.

scholarly journals Identification and Characterization of DM1 Patients by a New Diagnostic Certified Assay: Neuromuscular and Cardiac Assessments

2013 ◽  
Vol 2013 ◽  
pp. 1-6 ◽  
Author(s):  
Rea Valaperta ◽  
Valeria Sansone ◽  
Fortunata Lombardi ◽  
Chiara Verdelli ◽  
Alessio Colombo ◽  
...  
Keyword(s):  
Molecular Genetic ◽  
Clinical Manifestations ◽  
Molecular Method ◽  
High Reproducibility ◽  
Pathological Mechanism ◽  
Genetic Techniques ◽  
Identification And Characterization ◽  
Very High ◽  
Unambiguous Interpretation

The expansion of the specific trinucleotide sequence, [CTG], is the molecular pathological mechanism responsible for the clinical manifestations of DM1. Many studies have described different molecular genetic techniques to detect DM1, but as yet there is no data on the analytical performances of techniques used so far in this disease. We therefore developed and validated a molecular method, “Myotonic Dystrophy SB kit,” to better characterize our DM1 population. 113 patients were examined: 20 DM1-positive, 11 DM1/DM2-negative, and13 DM1-negative/DM2-positive, who had a previous molecular diagnosis, while 69 were new cases. This assay correctly identified 113/113 patients, and all were confirmed by different homemade assays. Comparative analysis revealed that the sensitivity and the specificity of the new kit were very high (>99%). Same results were obtained using several extraction procedures and different concentrations of DNA. The distribution of pathologic alleles showed a prevalence of the “classical” form, while of the 96 nonexpanded alleles 19 different allelic types were observed. Cardiac and neuromuscular parameters were used to clinically characterize our patients and support the new genetic analysis. Our findings suggest that this assay appears to be a very robust and reliable molecular test, showing high reproducibility and giving an unambiguous interpretation of results.

scholarly journals Cobalamin (vitamin B12) biosynthesis: identification and characterization of a Bacillus megaterium cobI operon

1998 ◽  
Vol 335 (1) ◽  
pp. 159-166 ◽  
Author(s):  
Evelyne RAUX ◽  
Anne LANOIS ◽  
Martin J. WARREN ◽  
Alain RAMBACH ◽  
Claude THERMES
Keyword(s):  
Bacillus Megaterium ◽  
Genomic Library ◽  
Early Stage ◽  
Open Reading Frames ◽  
Biosynthetic Genes ◽  
Pseudomonas Denitrificans ◽  
Dna Fragment ◽  
Identification And Characterization ◽  
Synechocystis Sp

A 16 kb DNA fragment has been isolated from a Bacillus megaterium genomic library and fully sequenced. The fragment contains 15 open reading frames, 14 of which are thought to constitute a B. megaterium cobalamin biosynthetic (cob) operon. Within the operon, 11 genes display similarity to previously identified Salmonella typhimurium cobalamin biosynthetic genes (cbiH60, -J, -C, -D, -ET, -L, -F, -G, -A, cysGA and btuR), whereas three do not (cbiW, -X and -Y). The genes of the B. megaterium cob operon were compared with the cobalamin biosynthetic genes of Pseudomonas denitrificans, Methanococcus jannaschii and Synechocystis sp. Taking into account the presence of cbiD and cbiG, the absence of a cobF, cobG and cobN, -S and -T, it was concluded that B. megaterium, M. jannaschii and Synechocystis sp., like S. typhimurium, synthesize cobalamin by an anaerobic pathway, in which cobalt is added at an early stage and molecular oxygen is not required.

scholarly journals Design and Optimization of a Rapid, Multiplex miRNA Assay without Washing Steps

Proceedings ◽  
2020 ◽  
Vol 60 (1) ◽  
pp. 32
Author(s):  
Giuliano Zanchetta ◽  
Thomas Carzaniga ◽  
Luka Vanjur ◽  
Luca Casiraghi ◽  
Giovanni Tagliabue ◽  
...  
Keyword(s):  
Early Stage ◽  
Optical Biosensor ◽  
Label Free ◽  
Time Intervals ◽  
One Pot ◽  
Design And Optimization ◽  
Individual Interaction ◽  
Equilibrium And Kinetics ◽  
Kinetics Of

MicroRNAs are widely studied as circulating biomarkers for early stage diagnosis of several diseases, but the procedures for their detection and quantification are currently complex and time consuming. We demonstrate a rapid, multiplex, one-pot detection method based on two-step amplification of the signal measured by a recent label-free optical biosensor, Reflective Phantom Interface (RPI). The specific capture with surface DNA probes is combined with mass amplification by an antibody targeting DNA–RNA hybrids and polyclonal secondary antibody, all performed without washing steps. Through this method, we achieved linear, sub-pM quantification of different miRNAs in 1.5 h. The RPI enabled the characterization of equilibrium and kinetics of each individual interaction involved in this multi-step process, which allowed us to model and optimize the relative concentrations and the time intervals of the assay.

scholarly journals Network Hamiltonian models reveal pathways to amyloid fibril formation

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Yue Yu ◽  
Gianmarc Grazioli ◽  
Megha H. Unhelkar ◽  
Rachel W. Martin ◽  
Carter T. Butts
Keyword(s):  
Amyloid Fibril ◽  
Prion Diseases ◽  
Data Bank ◽  
Fibril Formation ◽  
Toxic Species ◽  
Hamiltonian Models ◽  
Amyloid Fibril Formation ◽  
Wide Range ◽  
Kinetics Of

Abstract Amyloid fibril formation is central to the etiology of a wide range of serious human diseases, such as Alzheimer’s disease and prion diseases. Despite an ever growing collection of amyloid fibril structures found in the Protein Data Bank (PDB) and numerous clinical trials, therapeutic strategies remain elusive. One contributing factor to the lack of progress on this challenging problem is incomplete understanding of the mechanisms by which these locally ordered protein aggregates self-assemble in solution. Many current models of amyloid deposition diseases posit that the most toxic species are oligomers that form either along the pathway to forming fibrils or in competition with their formation, making it even more critical to understand the kinetics of fibrillization. A recently introduced topological model for aggregation based on network Hamiltonians is capable of recapitulating the entire process of amyloid fibril formation, beginning with thousands of free monomers and ending with kinetically accessible and thermodynamically stable amyloid fibril structures. The model can be parameterized to match the five topological classes encompassing all amyloid fibril structures so far discovered in the PDB. This paper introduces a set of network statistical and topological metrics for quantitative analysis and characterization of the fibrillization mechanisms predicted by the network Hamiltonian model. The results not only provide insight into different mechanisms leading to similar fibril structures, but also offer targets for future experimental exploration into the mechanisms by which fibrils form.

A Novel SCN5A Variant Associated with Abnormal Repolarization, Atrial Fibrillation, and Reversible Cardiomyopathy

Cardiology ◽  
2018 ◽  
Vol 140 (1) ◽  
pp. 8-13 ◽  
Author(s):  
Kim Boddum ◽  
Arnela Saljic ◽  
Thomas Jespersen ◽  
Alex Hørby Christensen
Keyword(s):  
Atrial Fibrillation ◽  
Clinical Manifestations ◽  
Persistent Atrial Fibrillation ◽  
Inactivation Kinetics ◽  
Loss Of Function ◽  
Na Current ◽  
Structural Abnormalities ◽  
Kinetics Of

A variety of life-threating arrhythmias are caused by mutations in the cardiac voltage-gated sodium channel encoded by the SCN5A gene. In this study, we report a novel loss-of-function SCN5A variant, p.Ile1343Val (c.4027A>G), identified in a 42-year-old proband who presented with an unusual ECG with abnormal repolarization with biphasic T-waves in anteroseptal leads, persistent atrial fibrillation (AF), intermittent left bundle branch block (LBBB), and reversible cardiomyopathy. The patient did not meet the diagnostic criteria for Brugada syndrome, long QT syndrome, or any other known SCN5A-associated phenotype. Characterization of the biophysical properties of the variant by in vitro patch clamp experiments revealed a reduced Na+ current with no effect on the inactivation kinetics of the channel. This loss-of-function of Na+ current could explain the intermittent LBBB as well as the AF. In conclusion, we describe a unique combination of electrical and structural abnormalities associated with a novel SCN5A variant. Our findings broaden the spectrum of cardiac phenotypes associated with SCN5A channelopathy, underlining the complex clinical manifestations of genetic variations within this gene.

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