Expression, oxidative refolding, and characterization of six-histidine-tagged recombinant human LECT2, a 16-kDa chemotactic protein with three disulfide bonds

2003 ◽  
Vol 27 (2) ◽  
pp. 272-278 ◽  
Author(s):  
Mie Ito ◽  
Koji Nagata ◽  
Yusuke Kato ◽  
Yoshifumi Oda ◽  
Satoshi Yamagoe ◽  
...  
Keyword(s):  
Disulfide Bonds ◽  
Oxidative Refolding ◽  
Chemotactic Protein

Characterization of disulfide bonds in recombinant proteins: reduced human interleukin 2 in inclusion bodies and its oxidative refolding

Biochemistry ◽  
1987 ◽  
Vol 26 (11) ◽  
pp. 3129-3134 ◽  
Author(s):  
Takashi Tsuji ◽  
Ryusuke Nakagawa ◽  
Nobuyuki Sugimoto ◽  
Kenichi Fukuhara
Keyword(s):  
Recombinant Proteins ◽  
Disulfide Bonds ◽  
Inclusion Bodies ◽  
Interleukin 2 ◽  
Oxidative Refolding

Morphologic and Molecular Characterization of Salmonella typhimurium Infection in Neonatal Calves

2002 ◽  
Vol 39 (2) ◽  
pp. 200-215 ◽  
Author(s):  
R. L. Santos ◽  
S. Zhang ◽  
R. M. Tsolis ◽  
A. J. Bäumler ◽  
L. G. Adams
Keyword(s):  
Salmonella Typhimurium ◽  
Rna Extraction ◽  
Specific Cell ◽  
Loop Model ◽  
Ileal Loop ◽  
Chemotactic Protein ◽  
Tnf Α ◽  
Holstein Calves ◽  
Neonatal Calves

The host response to Salmonella plays a major role in the outcome of infection. The present study was undertaken to further characterize Salmonella typhimurium infection in neonatal calves at both the morphologic and the molecular level using the ligated ileal loop model. Eight 4–5-week-old male Holstein calves underwent laparotomy, and loops were prepared in the ileum. The loops were either inoculated with an S. typhimurium strain pathogenic for cattle or injected with sterile LB broth as control. Samples for histology, transmission and scanning electron microscopy, and RNA extraction were collected at various time points between 5 minutes and 12 hours postinfection. Invasion of both M cells and enterocytes began at 15 minutes postinfection. No specific cell type was the main target for invasion. Intracellular bacteria were observed in the lamina propria after 1 hour postinfection. A severe acute neutrophilic response was associated with invasion of the Peyer's patches. Upregulated expression of CXC chemokines (interleukin [IL]-8, growth-related oncogenes, [GRO] α and γ, and granulocyte chemotactic protein [GCP]2) was detected by reverse transcription polymerase chain reaction beginning at 1 hour postinfection. Expression of proinflammatory (IL-1β, IL-18, and tumor necrosis factor [TNF]α) and anti-inflammatory (IL-10, IL-1Ra, and IL-4) cytokines was also assessed. A marked increase in expression of IL-1β was observed, whereas the profile of expression of IL-18 and TNFα did not change after infection. Upregulation of IL-1Ra and IL-4 but not of IL-10 was observed. These findings indicate that infection of bovine ligated ileal loops with S. typhimurium results in an acute neutrophilic inflammatory response that is associated with the upregulation of CXC chemokines (IL-8, GROα and γ, and GCP2), IL-1β, IL-1Ra, and IL-4.

scholarly journals Production and Characterization of Keratin/Tragacanth Gum Nanohydrogels for Drug Delivery in Medical Textiles

2021 ◽  
Vol 8 ◽  
Author(s):  
Nazanin Mansouri Shirazi ◽  
Niloofar Eslahi ◽  
Adeleh Gholipour-Kanani
Keyword(s):  
Disulfide Bonds ◽  
Antibacterial Properties ◽  
Antimicrobial Properties ◽  
Spherical Shape ◽  
Herbal Extract ◽  
Minimum Size ◽  
Mechanical Resistance ◽  
Tragacanth Gum ◽  
Medical Textiles

Keratin protein has been applied for biomedical applications due to its biocompatibility, biodegradability, mechanical resistance, and bioavailability. Tragacanth gum (TG) as a polysaccharide-based biopolymer has wound healing and antimicrobial properties. In this study, keratin was extracted from protein-based chicken feather by using reduction hydrolysis (sodium sulfide), and nanogels of keratin and TG composites at different ratios were produced by using the chemical cross-linking method. Then, cinnamon (5 and 10%) as an antibacterial herbal extract was added to the nanogels and coated on cotton fabric. The morphology and size of the composite nanogels, chemical structure, biological, and antibacterial properties were evaluated. According to DLS results, TGK2:1 (ratio of TG to keratin = 2:1) had the minimum size (80 nm) and PDI (0.1), and therefore, this sample was chosen as the optimum one. FESEM and TEM images showed the semi-spherical shape of the produced nanogels. FTIR spectra revealed the possible hydrogen bonding between the components, and the formation of disulfide bonds after the addition of hydrogen peroxide was confirmed by XPS. After loading cinnamon into the nanogels, an increase in size was observed from 80 nm for free-nanogel to 85 and 105 nm for 5 and 10% extract-loaded nanogels, respectively. Besides, more cinnamon was released from the treated fabrics by increasing time and cinnamon concentration. The antibacterial test exhibited good antibacterial properties against both Gram-positive and Gram-negative bacteria. Finally, MTT assay approved the biocompatibility of the produced nanogels for potential use in medical textiles.

scholarly journals Characterization of Intramolecular Disulfide Bonds and Secondary Modifications of the Glycoprotein from Viral Hemorrhagic Septicemia Virus, a Fish Rhabdovirus

1998 ◽  
Vol 72 (12) ◽  
pp. 10189-10196 ◽  
Author(s):  
Katja Einer-Jensen ◽  
Thomas N. Krogh ◽  
Peter Roepstorff ◽  
Niels Lorenzen
Keyword(s):  
G Protein ◽  
Disulfide Bonds ◽  
Chemical Reduction ◽  
Viral Hemorrhagic Septicemia Virus ◽  
Spectrometric Analysis ◽  
Glycan Structure ◽  
Viral Hemorrhagic Septicemia ◽  
Transmembrane Glycoprotein ◽  
Before And After

ABSTRACT Viral hemorrhagic septicemia virus (VHSV) infections cause high losses in cultured rainbow trout in Europe. Attempts to produce a recombinant vaccine based on the transmembrane glycoprotein (G protein) have indicated that proper folding is important for the antigenicity and immunogenicity of the protein. The present study was initiated to identify the disulfide bonds and other structural aspects relevant to vaccine design. The N-terminal amino acid residue was identified as being a pyroglutamic acid, corresponding to Gln21 of the primary transcript. Peptides from endoproteinase-degraded G protein were analyzed by mass spectrometry before and after chemical reduction, and six disulfide bonds were identified: Cys29-Cys339, Cys44-Cys295, Cys90-Cys132, Cys172-Cys177, Cys195-Cys265, and Cys231-Cys236. Mass spectrometric analysis in combination with glycosidases allowed characterization of the glycan structure of the G protein. Three of four predicted N-linked oligosaccharides were found to be predominantly biantennary complex-type structures. Furthermore, an O-linked glycan near the N terminus was identified. Alignment of the VHSV G protein with five other rhabdovirus G proteins indicates that eight cysteine residues are situated at conserved positions. This finding suggests that there might be some common disulfide bonding pattern among the six rhabdoviruses.

scholarly journals In-Depth Characterization of Protein Disulfide Bonds by Online Liquid Chromatography-Electrochemistry-Mass Spectrometry

2015 ◽  
Vol 27 (1) ◽  
pp. 50-58 ◽  
Author(s):  
Linda Switzar ◽  
Simone Nicolardi ◽  
Julie W. Rutten ◽  
Saskia A. J. Lesnik Oberstein ◽  
Annemieke Aartsma-Rus ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Disulfide Bonds ◽  
Protein Disulfide Bonds ◽  
Protein Disulfide

scholarly journals Characterization of KCNQ1 atrial fibrillation mutations reveals distinct dependence on KCNE1

2012 ◽  
Vol 139 (2) ◽  
pp. 135-144 ◽  
Author(s):  
Priscilla J. Chan ◽  
Jeremiah D. Osteen ◽  
Dazhi Xiong ◽  
Michael S. Bohnen ◽  
Darshan Doshi ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Disulfide Bonds ◽  
Functional Dependence ◽  
Voltage Dependent ◽  
Familial Atrial Fibrillation ◽  
Membrane Spanning ◽  
Mutant Phenotypes ◽  
Inherited Mutations ◽  
Iks Channel

The IKs potassium channel, critical to control of heart electrical activity, requires assembly of α (KCNQ1) and β (KCNE1) subunits. Inherited mutations in either IKs channel subunit are associated with cardiac arrhythmia syndromes. Two mutations (S140G and V141M) that cause familial atrial fibrillation (AF) are located on adjacent residues in the first membrane-spanning domain of KCNQ1, S1. These mutations impair the deactivation process, causing channels to appear constitutively open. Previous studies suggest that both mutant phenotypes require the presence of KCNE1. Here we found that despite the proximity of these two mutations in the primary protein structure, they display different functional dependence in the presence of KCNE1. In the absence of KCNE1, the S140G mutation, but not V141M, confers a pronounced slowing of channel deactivation and a hyperpolarizing shift in voltage-dependent activation. When coexpressed with KCNE1, both mutants deactivate significantly slower than wild-type KCNQ1/KCNE1 channels. The differential dependence on KCNE1 can be correlated with the physical proximity between these positions and KCNE1 as shown by disulfide cross-linking studies: V141C forms disulfide bonds with cysteine-substituted KCNE1 residues, whereas S140C does not. These results further our understanding of the structural relationship between KCNE1 and KCNQ1 subunits in the IKs channel, and provide mechanisms for understanding the effects on channel deactivation underlying these two atrial fibrillation mutations.

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