New insights about the self-aggregation of benzoporphyrin derivatives: A theoretical and experimental investigation

2018 ◽  
Vol 22 (04) ◽  
pp. 342-354 ◽  
Author(s):  
Gabriel B. Cesar ◽  
Diogo S. Pellosi ◽  
Douglas Vanzin ◽  
Renato S. Gonçalves ◽  
Wilker Caetano ◽  
...  
Keyword(s):  
Sodium Dodecyl ◽  
Ester Group ◽  
The Self ◽  
Van Der Waals Interactions ◽  
Molecular Volume ◽  
Aggregation Process ◽  
New Development ◽  
Self Aggregation

The self-aggregation process and its overcoming remain a barrier for the employment of many molecules as photosensitizers (PS) in photodynamic therapy (PDT). The B-ring isomer co-produced during the synthesis of the Verteporfin[Formula: see text] (A-ring) is an example. Although both isomers possess similar in vitro/in vivo efficiency, the strong and not well-understood self-aggregation process of the B-ring derivative impairs its clinical use. This paper reports the use of theoretical calculus and its correlation with experimental analysis to find the main differences between the A and B-ring isomers. For that purpose, micelles of Pluronic[Formula: see text] P-123 and Sodium Dodecyl Sulfate were chosen as simple membrane models and possible drug delivery system, as in the case of P-123. At physiological pH, the main reason for the high self-aggregation tendency is associated with the higher (22%) molecular volume of the B ring, which increases the van der Waals interactions. However, at mildly acidic conditions, the B ring possesses a shallow dihedral angle between the methyl ester group and the tetrapyrrolic macrocycle that favors the approach of units in the aggregate. These discrepancies directly affect the binding and stability of the isomers in the micelles. However, P-123 micelles were able to readily incorporate and monomerize/stabilize both PS over long periods. NOESY experiments confirmed a deep location of both PS inside P-123 micelles, which justifies their efficiency in preventing the self-aggregation process. These findings may substantiate new studies involving the marginalized B-ring isomers and encourage new development in formulations for their use in PDT.

scholarly journals Interaction of Aβ42 with Membranes Triggers the Self-Assembly into Oligomers

2020 ◽  
Vol 21 (3) ◽  
pp. 1129 ◽  
Author(s):  
Siddhartha Banerjee ◽  
Mohtadin Hashemi ◽  
Karen Zagorski ◽  
Yuri L. Lyubchenko
Keyword(s):  
Self Assembly ◽  
Membrane Surface ◽  
Amyloid Β ◽  
The Self ◽  
Bulk Solution ◽  
Aggregation Process ◽  
Physiological Concentration ◽  
Dynamics Simulations

The self-assembly of amyloid β (Aβ) proteins into oligomers is the major pathogenic event leading to Alzheimer’s disease (AD). Typical in vitro experiments require high protein concentrations, whereas the physiological concentration of Aβ is in the picomolar to low nanomolar range. This complicates the translation of results obtained in vitro to understanding the aggregation process in vivo. Here, we demonstrate that Aβ42 self-assembles into aggregates on membrane bilayers at low nanomolar concentrations - a pathway in which the membrane plays the role of a catalyst. Additionally, physiological ionic conditions (150 mM NaCl) significantly enhance on-membrane aggregation, leading to the rapid formation of oligomers. The self-assembly process is reversible, so assembled aggregates can dissociate from the membrane surface into the bulk solution to further participate in the aggregation process. Molecular dynamics simulations demonstrate that the transient membrane-Aβ interaction dramatically changes the protein conformation, facilitating the assembly of dimers. The results indicate peptide–membrane interaction is the critical step towards oligomer formation at physiologically low protein concentrations.

scholarly journals Effective decellularisation of human saphenous veins for biocompatible arterial tissue engineering applications: Bench optimisation and feasibility in vivo testing

2021 ◽  
Vol 12 ◽  
pp. 204173142098752
Author(s):  
Nadiah S Sulaiman ◽  
Andrew R Bond ◽  
Vito D Bruno ◽  
John Joseph ◽  
Jason L Johnson ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Mechanical Strength ◽  
Vascular Tissue ◽  
Sodium Dodecyl ◽  
Host Cells ◽  
Replacement Model ◽  
In Vivo Testing ◽  
Vascular Scaffolds

Human saphenous vein (hSV) and synthetic grafts are commonly used conduits in vascular grafting, despite high failure rates. Decellularising hSVs (D-hSVs) to produce vascular scaffolds might be an effective alternative. We assessed the effectiveness of a detergent-based method using 0% to 1% sodium dodecyl sulphate (SDS) to decellularise hSV. Decellularisation effectiveness was measured in vitro by nuclear counting, DNA content, residual cell viability, extracellular matrix integrity and mechanical strength. Cytotoxicity was assessed on human and porcine cells. The most effective SDS concentration was used to prepare D-hSV grafts that underwent preliminary in vivo testing using a porcine carotid artery replacement model. Effective decellularisation was achieved with 0.01% SDS, and D-hSVs were biocompatible after seeding. In vivo xeno-transplantation confirmed excellent mechanical strength and biocompatibility with recruitment of host cells without mechanical failure, and a 50% patency rate at 4-weeks. We have developed a simple biocompatible methodology to effectively decellularise hSVs. This could enhance vascular tissue engineering toward future clinical applications.

scholarly journals Antimicrobial Photodynamic Inactivation Inhibits Candida albicans Virulence Factors and ReducesIn VivoPathogenicity

2012 ◽  
Vol 57 (1) ◽  
pp. 445-451 ◽  
Author(s):  
Ilka Tiemy Kato ◽  
Renato Araujo Prates ◽  
Caetano Padial Sabino ◽  
Beth Burgwyn Fuchs ◽  
George P. Tegos ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Virulence Factors ◽  
Sodium Dodecyl ◽  
Systemic Infection ◽  
Tube Formation ◽  
Photodynamic Inactivation ◽  
Content Type ◽  
Daughter Cells

ABSTRACTThe objective of this study was to evaluate whetherCandida albicansexhibits altered pathogenicity characteristics following sublethal antimicrobial photodynamic inactivation (APDI) and if such alterations are maintained in the daughter cells.C. albicanswas exposed to sublethal APDI by using methylene blue (MB) as a photosensitizer (0.05 mM) combined with a GaAlAs diode laser (λ 660 nm, 75 mW/cm2, 9 to 27 J/cm2).In vitro, we evaluated APDI effects onC. albicansgrowth, germ tube formation, sensitivity to oxidative and osmotic stress, cell wall integrity, and fluconazole susceptibility.In vivo, we evaluatedC. albicanspathogenicity with a mouse model of systemic infection. Animal survival was evaluated daily. Sublethal MB-mediated APDI reduced the growth rate and the ability ofC. albicansto form germ tubes compared to untreated cells (P< 0.05). Survival of mice systemically infected withC. albicanspretreated with APDI was significantly increased compared to mice infected with untreated yeast (P< 0.05). APDI increasedC. albicanssensitivity to sodium dodecyl sulfate, caffeine, and hydrogen peroxide. The MIC for fluconazole forC. albicanswas also reduced following sublethal MB-mediated APDI. However, none of those pathogenic parameters was altered in daughter cells ofC. albicanssubmitted to APDI. These data suggest that APDI may inhibit virulence factors and reducein vivopathogenicity ofC. albicans. The absence of alterations in daughter cells indicates that APDI effects are transitory. The MIC reduction for fluconazole following APDI suggests that this antifungal could be combined with APDI to treatC. albicansinfections.

scholarly journals Protein synthesis in chloroplasts. Characteristics and products of protein synthesis in vitro in etioplasts and developing chloroplasts from pea leaves

1975 ◽  
Vol 146 (3) ◽  
pp. 675-685 ◽  
Author(s):  
S G Siddell ◽  
R J Ellis
Keyword(s):  
Energy Source ◽  
Protein Synthesis ◽  
Large Subunit ◽  
Sodium Dodecyl ◽  
Supernatant Fraction ◽  
Pisum Sativum L ◽  
Fraction I ◽  
Plastid Ribosomes

The function of plastid ribosomes in pea (Pisum sativum L.) was investigated by characterizing the products of protein synthesis in vitro in plastids isolated at different stages during the transition from etioplast to chloroplast. Etioplasts and plastids isolated after 24, 48 and 96h of greening in continuous white light, use added ATP to incorporate labelled amino acids into protein. Plastids isolated from greening leaves can also use light as the source of energy for protein synthesis. The labelled polypeptides synthesized in isolated plastids were analysed by electrophoresis in sodium dodecyl sulphate-ureapolyacrylamide gels. Six polypeptides are synthesized in etioplasts with ATP as energy source. Only one of these polypeptides is present in a 150 000g supernatant fraction. This polypeptide has been identified as the large subunit of Fraction I protein (3-phospho-D-glycerate carboxylyase EC 4.1.1.39) by comparing the tryptic ‘map’ of its L-(35S)methionine-labelled peptides with the tryptic ‘map’ of large subunit peptides from Fraction I labelled with L-(35S)methionine in vivo. The same gel pattern of six polypeptides is seen when plastids isolated from greening leaves are incubated with either added ATP or light as the energy source. However, the rates of synthesis of particular polypeptides are different in plastids isolated at different stages of the etioplast to chloroplast transition. The results support the idea that plastid ribosomes synthesize only a small number of proteins, and that the number and molecular weight of these proteins does not alter during the formation of chloroplasts from etioplasts.

UHF-1, a factor required for maximal transcription of early and late sea urchin histone H4 genes: analysis of promoter-binding sites

1991 ◽  
Vol 11 (2) ◽  
pp. 1048-1061
Author(s):  
I J Lee ◽  
L Tung ◽  
D A Bumcrot ◽  
E S Weinberg
Keyword(s):  
Binding Site ◽  
Sea Urchin ◽  
Transcriptional Start Site ◽  
Sodium Dodecyl ◽  
Nuclear Extract ◽  
Dnase I ◽  
Histone H4 ◽  
Band Shift

A protein, denoted UHF-1, was found to bind upstream of the transcriptional start site of both the early and late H4 (EH4 and LH4) histone genes of the sea urchin Strongylocentrotus purpuratus. A nuclear extract from hatching blastulae contained proteins that bind to EH4 and LH4 promoter fragments in a band shift assay and produced sharp DNase I footprints upstream of the EH4 gene (from -133 to -106) and the LH4 gene (from -94 to -66). DNase I footprinting performed in the presence of EH4 and LH4 promoter competitor DNAs indicated that UHF-1 binds more strongly to the EH4 site. A sequence match of 11 of 13 nucleotides was found within the two footprinted regions: [sequence: see text]. Methylation interference and footprinting experiments showed that UHF-1 bound to the two sites somewhat differently. DNA-protein UV cross-linking studies indicated that UHF-1 has an electrophoretic mobility on sodium dodecyl sulfate-acrylamide gels of approximately 85 kDa and suggested that additional proteins, specific to each promoter, bind to each site. In vitro and in vivo assays were used to demonstrate that the UHF-1-binding site is essential for maximal transcription of the H4 genes. Deletion of the EH4 footprinted region resulted in a 3-fold decrease in transcription in a nuclear extract and a 2.6-fold decrease in expression in morulae from templates that had been injected into eggs. In the latter case, deletion of the binding site did not grossly disrupt the temporal program of expression from the injected EH4 genes. LH4 templates containing a 10-bp deletion in the consensus region or base substitutions in the footprinted region were transcribed at 14 to 58% of the level of the wild-type LH4 template. UHF-1 is therefore essential for maximal expression of the early and late H4 genes.

scholarly journals Isolation and Characterization of Two Proteins fromMoraxella catarrhalis That Bear a Common Epitope

1998 ◽  
Vol 66 (9) ◽  
pp. 4374-4381 ◽  
Author(s):  
John C. McMichael ◽  
Michael J. Fiske ◽  
Ross A. Fredenburg ◽  
Deb N. Chakravarti ◽  
Karl R. VanDerMeid ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Bacterial Attachment ◽  
Vaccine Candidates ◽  
Isolation And Characterization ◽  
Sds Page

ABSTRACT The UspA1 and UspA2 proteins of Moraxella catarrhalisare potential vaccine candidates for preventing disease caused by this organism. We have characterized both proteins and evaluated their vaccine potential using both in vitro and in vivo assays. Both proteins were purified from the O35E isolate by Triton X-100 extraction, followed by ion-exchange and hydroxyapatite chromatography. Analysis of the sequences of internal peptides, prepared by enzymatic and chemical cleavage of the proteins, revealed that UspA1 and UspA2 exhibited distinct structural differences but shared a common sequence including an epitope recognized by the monoclonal antibody 17C7. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), purified UspA1 exhibited a molecular weight of approximately 350,000 when unheated and a molecular weight of 100,000 after being heated for 10 min at 100°C. In contrast, purified UspA2 exhibited an apparent molecular weight of 240,000 by SDS-PAGE that did not change with the length of time of heating. Their sizes as determined by gel filtration were 1,150,000 and 830,000 for UspA1 and UspA2, respectively. Preliminary results indicate the proteins have separate functions in bacterial pathogenesis. Purified UspA1 was found to bind HEp-2 cells, and sera against UspA1, but not against UspA2, blocked binding of the O35E isolate to the HEp-2 cells. UspA1 also bound fibronectin and appears to have a role in bacterial attachment. Purified UspA2, however, did not bind fibronectin but had an affinity for vitronectin. Both proteins elicited bactericidal antibodies in mice to homologous and heterologous disease isolates. Finally, mice immunized with each of the proteins, followed by pulmonary challenge with either the homologous or a heterologous isolate, cleared the bacteria more rapidly than mock-immunized mice. These results suggest that UspA1 and UspA2 serve different virulence functions and that both are promising vaccine candidates.

scholarly journals The Clinical Application of Porous Tantalum and Its New Development for Bone Tissue Engineering

2021 ◽  
Author(s):  
Gan Huang ◽  
Shu-Ting Pan ◽  
Jia-Xuan Qiu
Keyword(s):  
Clinical Application ◽  
Bone Ingrowth ◽  
Primary Stability ◽  
High Porosity ◽  
Marginal Bone Loss ◽  
Clinical Value ◽  
Porous Tantalum ◽  
New Development

Porous tantalum (Ta) is a promising biomaterial and has been applied in orthopedics and dentistry for nearly two decades. The high porosity and interconnected pore structure of porous Ta promise fine bone ingrowth and new bone formation within the inner space, which further guarantee rapid osteointegration and bone-implant stability in long term. Porous Ta has high wettability and surface energy that can facilitate adherence, proliferation and mineralization of osteoblasts. Meanwhile, low elastic modulus and high friction coefficient of porous Ta can effectively avoid stress shield effect, minimize marginal bone loss and ensure primary stability. Accordingly, the satisfactory clinical application of porous Ta based implants or prostheses are mainly derived from its excellent biological and mechanical properties. With the advent of additive manufacturing, personalized porous Ta based implants or prostheses have shown their clinical value in the treatment of individual patient who need specially designed implant or prosthesis. In addition, many modification methods have been introduced to enhance the bioactivity and antibacterial property of porous Ta with promising in vitro and in vivo research results. In any case, choosing suitable patients is of great importance to guarantee surgical success after porous Ta insertion.

scholarly journals Characterization of proteins from the cytoskeleton of Giardia lamblia

1983 ◽  
Vol 59 (1) ◽  
pp. 81-103 ◽  
Author(s):  
R. Crossley ◽  
D.V. Holberton
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Giardia Lamblia ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Molecular Size ◽  
Molecular Weights

Proteins from the axonemes and disc cytoskeleton of Giardia lamblia have been examined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis. In addition to tubulin and the 30 X 10(3) molecular weight disc protein, at least 18 minor components copurify with the two major proteins in Triton-insoluble structures. The most prominent minor bands have the apparent molecular weights of 110 X 10(3), 95 X 10(3) and 81 X 10(3). Protein of 30 X 10(3) molecular weight accounts for about 20% of organelle protein on gels. In continuous 25 mM-Tris-glycine buffer it migrates mostly as a close-spaced doublet of polypeptides, which are here given the name giardins. Giardia tubulin and giardin have been purified by gel filtration chromatography in the presence of sodium dodecyl sulphate. Well-separated fractions were obtained that could be further characterized. Both proteins are heterogeneous when examined by isoelectric focusing. Five tubulin chains were detected by PAGE Blue 83 dye-binding after focusing in a broad-range ampholyte gel. Giardin is slightly less acidic than tubulin. On gels it splits into four major and four minor chains with isoelectric points in the pI range from 5.8 to 6.2. The amino acid composition of the giardin fraction has been determined, and compared to Giardia tubulin and a rat brain tubulin standard. Giardins are rich in helix-forming residues, particularly leucine. They have a low content of proline and glycine; therefore they may have extensive alpha-helical regions and be rod-shaped. As integral proteins of disc microribbons, giardins in vivo associate closely with tubulin. The properties of giardins indicate that in a number of respects - molecular size, charge, stoichiometry - their structural interaction with tubulin assemblies will be different from other tubulin-accessory protein copolymers studied in vitro.

Effect of microenvironment and cell-line type on carbohydrate-binding proteins of macrophage-like cells

1988 ◽  
Vol 66 (11) ◽  
pp. 1169-1176 ◽  
Author(s):  
Hans-Joachim Gabius ◽  
Katalin Vehmeyer
Keyword(s):  
Binding Proteins ◽  
Cultured Cells ◽  
Sodium Dodecyl ◽  
Carbohydrate Binding ◽  
Intercellular Interaction ◽  
Cell Extracts ◽  
Carbohydrate Binding Proteins ◽  
Sugar Receptors

The pattern of sugar inhibition of rosette formation, a model for intercellular interaction between cultured cells and glutaraldehyde-fixed, trypsinated rabbit erythrocytes, served to infer the presence of carbohydrate-binding proteins. This profile from cell extracts for the two murine macrophage-like cell lines, P388D1 and J774A.1, was comparatively analyzed by affinity chromatography on supports with immobilized carbohydrates (lactose, L-fucose, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, and maltose) or with the immobilized mannose-rich yeast glycoprotein mannan or fetuin-derived glycopeptides containing sialic acid residues. After elution with specific sugar in the absence of Ca2+ ions, the proteins were separated by sodium dodecyl sulfate – polyacrylamide slab gel electrophoresis. The composition of carbohydrate-binding proteins of the two lines clearly exhibited quantitative and qualitative differences. Moreover, the pattern of P388D1 cells was also demonstrated to change significantly in response to alterations in the conditions of the physiological environment. These alterations were imposed by in vitro growth, by subsequent in vivo growth in nude mice, and by re-adaptation of cells to culture after in vivo passage. Collectively, our observations and other physiological and biochemical reports on macrophage lectins indicate that the presence of sugar receptors with different specificities may be an indicator of macrophage differentiation, being reversibly modulated to a considerable extent by external factors, e.g., microenvironment. Extensive but selective alterations in this respect could play an important role in the control of recognition and effector mechanisms within diverse functions of macrophage subpopulations.

A family of related proteins is encoded by the major Drosophila heat shock gene family

1982 ◽  
Vol 2 (3) ◽  
pp. 286-292
Author(s):  
S C Wadsworth
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Heat Shock Protein ◽  
Sodium Dodecyl ◽  
Heat Shock Gene ◽  
Partial Digestion ◽  
Shock Gene ◽  
Shock Proteins

At least four proteins of 70,000 to 75,000 molecular weight (70-75K) were synthesized from mRNA which hybridized with a cloned heat shock gene previously shown to be localized to the 87A and 87C heat shock puff sites. These in vitro-synthesized proteins were indistinguishable from in vivo-synthesized heat shock-induced proteins when analyzed on sodium dodecyl sulfate-polyacrylamide gels. A comparison of the pattern of this group of proteins synthesized in vivo during a 5-min pulse or during continuous labeling indicates that the 72-75K proteins are probably not kinetic precursors to the major 70K heat shock protein. Partial digestion products generated with V8 protease indicated that the 70-75K heat shock proteins are closely related, but that there are clear differences between them. The partial digestion patterns obtained from heat shock proteins from the Kc cell line and from the Oregon R strain of Drosophila melanogaster are very similar. Genetic analysis of the patterns of 70-75K heat shock protein synthesis indicated that the genes encoding at least two of the three 72-75K heat shock proteins are located outside of the major 87A and 87C puff sites.

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