scholarly journals Applying Different Chemical Methods to Develop an Efficient Acellular Rat Uterine Tissue Matrix

2021 ◽  
Author(s):  
Masoomeh Masoomikarimi ◽  
Samira Rajaei ◽  
Farshid Noorbakhsh ◽  
Majid Salehi
Keyword(s):  
Blood Compatibility ◽  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Complete Removal ◽  
Dimensional Structure ◽  
Chemical Methods ◽  
Optimized Protocol ◽  
Intact Uterus ◽  
Tissue Matrix ◽  
Different Doses

Abstract Purpose: Decellularized uterine scaffold, as a new achievement in tissue engineering, permits recellularization and regeneration of uterine tissues and supports pregnancy in a fashion comparable to the intact uterus. The main purpose of this study was using of different chemical methods to introduce an optimized protocol for decellularization of rat uterus.Method: We decellularized rat uteruses by four different protocols using sodium dodecyl sulfate (SDS) and triton-X100 with different doses and time incubations.We characterized the scaffolds through histopathological staining, DNA quantification, MTT assay, Blood compatibility assay, Field Emission Scanning Electron Microscopy (FESEM), and biomechanical studies.Results: Histology assessment showed that only in protocol 4, cell residues were completely removed. Masson’s trichrome staining demonstrates that in protocol P3 collagen bundles were decreased; however, no damage in the collagen bundles was observed by other protocols. Cell viabilities indirect MTT assays of all protocols were significantly higher than the native samples. The RBC hemolysis percent in the presence of prepared scaffolds from all 4 protocols was less than 2%. The mechanical properties of none of the protocols were significantly different from the native sample. Conclusion: Protocol 4 which used freeze-thawing before using detergent, was introduced as the optimized protocol due to complete removal of cell residue, preservation of the three-dimensional structure, complete removal of detergents, and preservation of the mechanical property of the scaffolds.

scholarly journals Conformation and membrane interaction studies of the potent antimicrobial and anticancer peptide palustrin-Ca

2021 ◽  
Author(s):  
Patrick Brendan Timmons ◽  
Chandralal M Hewage
Keyword(s):  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Dynamics Simulation ◽  
Peptide Sequence ◽  
Helical Conformation ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Anticancer Activities ◽  
American Bullfrog ◽  
Α Helix

Palustrin-Ca (GFLDIIKDTGKEFAVKILNNLKCKLAGGCPP) is a host defense peptide with potent antimicrobial and anticancer activities, first isolated from the skin of the American bullfrog Lithobates catesbeianus. The peptide is 31 amino acid residues long, cationic and amphipathic. Two-dimensional NMR spectroscopy was employed to characterise its three-dimensional structure in a 50/50% water/2,2,2-trifluoroethanol-d3 mixture. The structure is defined by an α-helix that spans between Ile6-Ala26, and a cyclic disulphide bridged domain at the C-terminal end of the peptide sequence, between residues 23 and 29. A molecular dynamics simulation was employed to model the peptide's interactions with sodium dodecyl sulphate micelles, a widely used bacterial membrane-mimicking environment. Throughout the simulation, the peptide was found to maintain its α-helical conformation between residues Ile6-Ala26, while adopting a position parallel to the surface to micelle, which is energetically-favourable due to many hydrophobic and electrostatic contacts with the micelle.

Creating a tissue-specific microdispersed matrix from a decellularized porcine liver

2020 ◽  
Vol 4 ◽  
pp. 41-50
Author(s):  
A.D. Kirillova ◽  
◽  
Yu.B. Basok ◽  
A.E. Lazhko ◽  
A. M. Grigoryev ◽  
...  
Keyword(s):  
Sodium Dodecyl ◽  
Genetic Material ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Porcine Liver ◽  
Tissue Specific ◽  
Main Components ◽  
Triton X ◽  
Injectable Form

The main problem with decellularization of liver tissue as a tissue-specific matrix/scaffold in liver bioengineered structures is the need to maximize the preservation of the original three-dimensional structure of the tissue and the main components of its extracellular matrix (ECM) while removing cells and genetic material. The attempts to use the existing protocols for the decellularization of other tissues and organs have been unsuccessful. The aim of the work is to develop a method for creation of tissue-specific microdispersed matrix from decellularized porcine liver (TMM DLp). The protocol for decellularization of porcine liver (Lp) fragments has been developed based on the complex application of chemical (sodium dodecyl sulfate and Triton X-100), biochemical (DNase I), and physical (supercritical CO2) methods for treatment the initial tissue. As a result of the found optimal conditions for decellularization of Lp with subsequent cryomicronization of DLp, the injectable form of the microdispersed tissue-specific matrix was obtained, which represents DLp microparticles with the size of 100-200 microns with the residual amount of DNA no more than 10±1.5 ng/mg (less than 1.0%), with the preservation of the microstructure and basic composition of the liver ECM. According to the assessment of biocompatible properties in vitro, TMM DLp samples meet the criteria of biological safety for cytotoxicity and hemolytic activity.

scholarly journals Solution Structure of Acidocin B, a Circular Bacteriocin Produced by Lactobacillus acidophilus M46

2015 ◽  
Vol 81 (8) ◽  
pp. 2910-2918 ◽  
Author(s):  
Jeella Z. Acedo ◽  
Marco J. van Belkum ◽  
Christopher T. Lohans ◽  
Ryan T. McKay ◽  
Mark Miskolzie ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Lactobacillus Acidophilus ◽  
Solution Structure ◽  
Sequence Similarity ◽  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Peptide Sequence ◽  
Dimensional Structure ◽  
Content Type ◽  
Circular Bacteriocin

ABSTRACTAcidocin B, a bacteriocin produced byLactobacillus acidophilusM46, was originally reported to be a linear peptide composed of 59 amino acid residues. However, its high sequence similarity to gassericin A, a circular bacteriocin fromLactobacillus gasseriLA39, suggested that acidocin B might be circular as well. Acidocin B was purified from culture supernatant by a series of hydrophobic interaction chromatographic steps. Its circular nature was ascertained by matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry and tandem mass spectrometry (MS/MS) sequencing. The peptide sequence was found to consist of 58 amino acids with a molecular mass of 5,621.5 Da. The sequence of the acidocin B biosynthetic gene cluster was also determined and showed high nucleotide sequence similarity to that of gassericin A. The nuclear magnetic resonance (NMR) solution structure of acidocin B in sodium dodecyl sulfate micelles was elucidated, revealing that it is composed of four α-helices of similar length that are folded to form a compact, globular bundle with a central pore. This is a three-dimensional structure for a member of subgroup II circular bacteriocins, which are classified based on their isoelectric points of ∼7 or lower. Comparison of acidocin B with carnocyclin A, a subgroup I circular bacteriocin with four α-helices and a pI of 10, revealed differences in the overall folding. The observed variations could be attributed to inherent diversity in their physical properties, which also required the use of different solvent systems for three-dimensional structural elucidation.

SdsA polymorph isolation and improvement of their crystal quality using nonconventional crystallization techniques

2015 ◽  
Vol 48 (5) ◽  
pp. 1551-1559 ◽  
Author(s):  
Eugenio De la Mora ◽  
Edith Flores-Hernández ◽  
Jean Jakoncic ◽  
Vivian Stojanoff ◽  
Dritan Siliqi ◽  
...  
Keyword(s):  
Magnetic Field ◽  
Active Site ◽  
Crystal Packing ◽  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
X Ray Diffraction ◽  
Oligomeric State ◽  
X Ray ◽  
Internal Order

SdsA, a sodium dodecyl sulfate hydrolase, fromPseudomonas aeruginosawas crystallized in three different crystal polymorphs and their three-dimensional structure was determined. The different polymorphs present different crystal packing habits. One of the polymorphs suggests the existence of a tetramer, an oligomeric state not observed previously, while the crystal packing of the remaining two polymorphs obstructs the active site entrance but stabilizes flexible regions of the protein. Nonconventional crystallization methods that minimize convection, such as counterdiffusion in polyvinyl alcohol gel coupled with the influence of a 500 MHz (10.2 T) magnetic field, were necessary to isolate the poorest diffracting polymorph and increase its internal order to determine its structure by X-ray diffraction. The results obtained show the effectiveness of nonconventional crystallographic methods to isolate different crystal polymorphs.

scholarly journals A Major Outer Membrane Protein of Rahnella aquatilisFunctions as a Porin and Root Adhesin

1998 ◽  
Vol 180 (4) ◽  
pp. 909-913 ◽  
Author(s):  
Wafa Achouak ◽  
Jean-Marie Pages ◽  
Rene De Mot ◽  
Gerard Molle ◽  
Thierry Heulin
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Lipid Membrane ◽  
Single Channel ◽  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Major Outer Membrane Protein ◽  
Dimensional Structure ◽  
Terminal Portion

ABSTRACT A 38-kDa major outer membrane protein (OMP) was isolated from the nitrogen-fixing enterobacterium Rahnella aquatilis CF3. This protein exists as a stable trimer in the presence of 2% sodium dodecyl sulfate at temperatures below 60°C. Single channel experiments showed that this major OMP of R. aquatilis CF3 is able to form pores in the planar lipid membrane. Two oligonucleotides encoding the N-terminal portion of the 38-kDa OMP and C-terminal portion of OmpC were used to amplify the 38-kDa gene by PCR. The deduced amino acid sequence showed a strong homology withEscherichia coli, Klebsiella pneumoniae,Salmonella typhi, and Serratia marcescens OmpC sequences, except loops L6 and L7, which are postulated to be cell surface exposed. On the basis of the OmpF-PhoE three-dimensional structure, it seems likely that this 38-kDa organizes three 16-strand β-barrel subunits. The relationship between the structure and the double functionality of this protein as porin and as a root adhesin is discussed.

scholarly journals Crystallization of cytoplasmic actin in complex with deoxyribonuclease I

1985 ◽  
Vol 225 (2) ◽  
pp. 517-522 ◽  
Author(s):  
H G Mannherz ◽  
W Kabsch ◽  
D Suck ◽  
K Friebel ◽  
M Frimmer
Keyword(s):  
Smooth Muscle Actin ◽  
Polyacrylamide Gel Electrophoresis ◽  
Heavy Atom ◽  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Cysteine Residue ◽  
Dimensional Structure ◽  
Muscle Actin ◽  
Porcine Liver ◽  
Deoxyribonuclease I

Crystals of cytoplasmic (porcine liver) actin in complex with deoxyribonuclease I (DNAase I) were prepared for structural determination by X-ray-diffraction analysis. The crystallization of porcine liver actin-DNAase I complex is preceded by a brief treatment with immobilized trypsin, whereby a C-terminal tri- or di-peptide including cysteine-374 is removed from the actin without any noticeable degradation of both proteins as judged by sodium dodecyl-sulphate/polyacrylamide-gel electrophoresis. Analysis of the crystals obtained does not reveal any differences in the three-dimensional structure of porcine liver actin from its skeletal compartment at up to 0.6 nm resolution. However, in contrast with crystalline skeletal-muscle actin-DNAase I complex, heavy-atom substitution of crystals of porcine liver actin-DNAase I complex could not be achieved with methyl mercuriacetate. Evidence is presented that, in porcine liver actin, the N-terminal cysteine residue is not located at position no. 10, as in skeletal- and smooth-muscle actin, but most probably at position no. 17. Thus, because this site is covered by DNAase I, the cysteine becomes inaccessible to titration with 5,5′-dithiobis-(2-nitrobenzoic acid) after complex-formation with DNAase I.

scholarly journals Conformation and membrane interaction studies of the potent antimicrobial and anticancer peptide palustrin-Ca

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Patrick B. Timmons ◽  
Chandralal M. Hewage
Keyword(s):  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Dynamics Simulation ◽  
Peptide Sequence ◽  
Helical Conformation ◽  
Dimensional Structure ◽  
Amino Acid Residues ◽  
Anticancer Activities ◽  
Anticancer Peptide ◽  
American Bullfrog

AbstractPalustrin-Ca (GFLDIIKDTGKEFAVKILNNLKCKLAGGCPP) is a host defence peptide with potent antimicrobial and anticancer activities, first isolated from the skin of the American bullfrog Lithobates catesbeianus. The peptide is 31 amino acid residues long, cationic and amphipathic. Two-dimensional NMR spectroscopy was employed to characterise its three-dimensional structure in a 50/50% water/2,2,2-trifluoroethanol-$$d_{3}$$ d 3 mixture. The structure is defined by an $$\alpha$$ α -helix that spans between Ile$$^{6}$$ 6 -Ala$$^{26}$$ 26 , and a cyclic disulfide-bridged domain at the C-terminal end of the peptide sequence, between residues 23 and 29. A molecular dynamics simulation was employed to model the peptide’s interactions with sodium dodecyl sulfate micelles, a widely used bacterial membrane-mimicking environment. Throughout the simulation, the peptide was found to maintain its $$\alpha$$ α -helical conformation between residues Ile$$^{6}$$ 6 -Ala$$^{26}$$ 26 , while adopting a position parallel to the surface to micelle, which is energetically-favourable due to many hydrophobic and electrostatic contacts with the micelle.

Sign in / Sign up

Export Citation Format

Share Document