Browse
Submit Data
Databases
API
Help

Dataset Information

11 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Thiol and disulphide contents of hen ovalbumin. C-terminal sequence and location of disulphide bond.

ABSTRACT: 1. The thiol and disulphide contents of hen ovalbumin were investigated by p-chloromercuribenzoate titration, by determination of cysteic acid content after performic acid oxidation, by measurement of uptake of radioactive iodoacetic acid, and by assay of S-aminoethylcysteine after reaction with ethyleneimine. All results showed that ovalbumin had 6 half-cystine residues. Experiments with and without reducing agents demonstrated that there were 4 thiol groups and 1 disulphide bond. 2. A peptide containing equimolar amounts of S-carboxymethyl-cysteine, serine, valine and proline, but no lysine or arginine, was obtained by radioactive labelling of the cysteine residues with iodo[(14)C]acetic acid followed by electrophoretic and chromatographic separation of tryptic digests. It was concluded that the C-terminal sequence of ovalbumin is -Cys-Val-Ser-Pro. 3. The location of the disulphide bond was studied by using a double-labelling technique. It was shown that one end of the disulphide was located in this C-terminal peptide.

SUBMITTER: Fothergill LA

PROVIDER: S-EPMC1185399 | biostudies-other | 1970 Feb

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Disulphide bond inhibitors in 2 cancer cell lines

2013-11-06 | GSE41666 | GEO

Disulphide bond inhibitors in 2 cancer cell lines

Project description:To elucidate which ER cargo maturation processes may be oxygen dependent, we first reasoned that the transcriptional response may be tailored specifically to counteract the mechanism of the imposed stress. Therefore, we compared the transcriptional regulation of the ERome during anoxic conditions to ER stress caused by lack of glycosylation (tunicamycin) or chaperone inactivation by calcium depletion (thapsigargin) In two cancer cell lines (colon carcinoma HCT116, and hepatocellular carcinoma Hep G2) cells were given one of 4 treatments. 24 hours of 0.0% O2 (Anoxia), 1.5 µg/ml tunicamycin or 0.3 µM thapsigargin with a control 21% O2 (Normoxia). Cells were exposed to hypoxia and anoxia in H35 and H85 HypOxystations (Don Whitley Scientific). 3 biological replicates of each experiment were carried out. Total of 24 samples.

2013-11-06 | E-GEOD-41666 | biostudies-arrayexpress

Isomerization of the intersubunit disulphide-bond in Env controls retrovirus fusion.

Project description:The membrane fusion activity of murine leukaemia virus Env is carried by the transmembrane (TM) and controlled by the peripheral (SU) subunit. We show here that all Env subunits of the virus form disulphide-linked SU-TM complexes that can be disrupted by treatment with NP-40, heat or urea, or by Ca(2+) depletion. Thiol mapping indicated that these conditions induced isomerization of the disulphide-bond by activating a thiol group in a Cys-X-X-Cys (CXXC) motif in SU. This resulted in dissociation of SU from the virus. The active thiol was hidden in uninduced virus but became accessible for alkylation by either Ca(2+) depletion or receptor binding. The alkylation inhibited isomerization, virus fusion and infection. DTT treatment of alkylated Env resulted in cleavage of the SU-TM disulphide-bond and rescue of virus fusion. Further studies showed that virus fusion was specifically inhibited by high and enhanced by low concentrations of Ca(2+). These results suggest that Env is stabilized by Ca(2+) and that receptor binding triggers a cascade of reactions involving Ca(2+) removal, CXXC-thiol exposure, SU-TM disulphide-bond isomerization and SU dissociation, which lead to fusion activation.

| S-EPMC1271652 | biostudies-literature

Thiol-disulphide independent in-cell trapping for the identification of peroxiredoxin 2 interactors.

Project description:Hydrogen peroxide (H2O2) acts as a signalling molecule by oxidising cysteine thiols in proteins. Recent evidence has established a role for cytosolic peroxiredoxins in transmitting H2O2-based oxidation to a multitude of target proteins. Moreover, it is becoming clear that peroxiredoxins fulfil their function in organised microdomains, where not all interactors are covalently bound. However, most studies aimed at identifying peroxiredoxin interactors were based on methods that only detect covalently linked partners. Here, we explore the applicability of two thiol-disulphide independent in-cell trapping methodological approaches in combination with mass spectrometry for the identification of interaction partners of peroxiredoxin 2 (Prdx2). The first is biotin-dependent proximity-labelling (BioID) with a biotin ligase A (BirA*)-fused Prdx2, which has never been applied on redox-active proteins. The second is crosslinker co-immunoprecipitation with an N-terminally His-tagged Prdx2. During the initial characterisation of the tagged Prdx2 constructs, we found that the His-tag, but not BirA*, compromises the peroxidase and signalling activities of Prdx2. Further, the Prdx2 interactors identified with each approach showed little overlap. We therefore concluded that BioID is a more reliable method than crosslinker co-immunoprecipitation. After a stringent mass spec data filtering, BioID identified 13 interactors under elevated H2O2 conditions, including subunit five of the COP9 signalosome complex (CSN5). The Prdx2:CSN5 interaction was further confirmed in a proximity ligation assay. Taken together, our results demonstrate that BioID can be used as a method for the identification of interactors of Prdxs, and that caution should be exercised when interpreting protein-protein interaction results using tagged Prdxs.

| S-EPMC8346688 | biostudies-literature

A disulphide bond in the E2 enzyme Pex4p modulates ubiquitin-conjugating activity.

Project description:The ubiquitin-conjugating enzyme Pex4p together with its binding partner, the peroxisomal membrane protein Pex22p, co-ordinates cysteine-dependent ubiquitination of the cycling receptor protein Pex5p. Unusually for an ubiquitin-conjugating enzyme, Saccharomyces cerevisiae Pex4p can form a disulphide bond between the cysteine residues at positions 105 and 146. We found that mutating the disulphide forming cysteine residues in Pex4p to serines does not disturb the secondary structure of the protein but does reduce the in vitro activity of Pex4p. From the crystal structure of Pex4p C105S, C146S in complex with the soluble domain of Pex22p, we observe a narrowing of the active site cleft, caused by loss of the disulphide bond. This modification of the active site microenvironment is likely to restrict access of ubiquitin to the active site cysteine, modulating Pex4p activity. Finally, based on sequence and structural alignments, we have identified other ubiquitin-conjugating enzymes that may contain disulphide bonds.

| S-EPMC6505396 | biostudies-literature

Selective reduction of a disulphide bridge in hen ovotransferrin.

Project description:Brief treatment of iron-saturated hen ovotransferrin with dithiothreitol selectively cleaves the disulphide bridge between residues 478 and 671, which is in the C-terminal domain of the protein. The reduced alkylated protein is less stable than the native protein, and its iron-binding properties are different. A fluorescent derivative was prepared by coupling N-iodoacetyl-N'-(5-sulpho-1-naphthyl)ethylenediamine to the thiol groups.

| S-EPMC1145035 | biostudies-other

Dynamic nature of disulphide bond formation catalysts revealed by crystal structures of DsbB.

Project description:In the Escherichia coli system catalysing oxidative protein folding, disulphide bonds are generated by the cooperation of DsbB and ubiquinone and transferred to substrate proteins through DsbA. The structures solved so far for different forms of DsbB lack the Cys104-Cys130 initial-state disulphide that is directly donated to DsbA. Here, we report the 3.4 A crystal structure of a DsbB-Fab complex, in which DsbB has this principal disulphide. Its comparison with the updated structure of the DsbB-DsbA complex as well as with the recently reported NMR structure of a DsbB variant having the rearranged Cys41-Cys130 disulphide illuminated conformational transitions of DsbB induced by the binding and release of DsbA. Mutational studies revealed that the membrane-parallel short alpha-helix of DsbB has a key function in physiological electron flow, presumably by controlling the positioning of the Cys130-containing loop. These findings demonstrate that DsbB has developed the elaborate conformational dynamism to oxidize DsbA for continuous protein disulphide bond formation in the cell.

| S-EPMC2666032 | biostudies-literature

The location of the thiol groups of myosin that are protected by pyrophosphate against reaction with 2,4-dinitrophenyl -hydroxyethyl disulphide.

Project description:Myosin modified in the presence or in the absence of pyrophosphate by 2,4-dinitrophenyl beta-hydroxyethyl disulphide was treated with iodo[1-(14)C]acetamide. The residual Ca(2+)-stimulated adenosine triphosphatase (ATPase) activity of the modified myosin was different depending on the presence or absence of PP(i) during modification and the number of 2,4-dinitrophenyl beta-hydroxyethyl disulphide-modified thiol groups. The radioactivity incorporated into the light components of myosin correlated with the Ca(2+)-stimulated ATPase activity of the modified myosin and decreased with decreasing residual Ca(2+)-stimulated ATPase activity of the modified myosin. When native myosin was treated with low concentrations of iodo[1-(14)C]acetamide the residual Ca(2+)-stimulated ATPase activity of carboxyamidomethylated myosin was high and the radioactivity incorporated into the light components of myosin was negligible. The thiol groups of the light components of myosin are essential to preserve the ATPase activity of the protein and are close to the pyrophosphate-binding sites.

| S-EPMC1178049 | biostudies-other

Vitamin D status is independently associated with plasma glutathione and cysteine thiol/disulphide redox status in adults.

Project description:Redox status and inflammation are important in the pathophysiology of numerous chronic diseases. Epidemiological studies have linked vitamin D status to a number of chronic diseases. We aimed to examine the relationships between serum 25-hydroxyvitamin D [25(OH)D] and circulating thiol/disulphide redox status and biomarkers of inflammation.This was a cross-sectional study of N = 693 adults (449 females, 244 males) in an apparently healthy, working cohort in Atlanta, GA. Plasma glutathione (GSH), cysteine (Cys) and their associated disulphides were determined with high-performance liquid chromatography, and their redox potentials (Eh GSSG and Eh CySS) were calculated using the Nernst equation. Serum inflammatory markers included interleukin-6 (IL-6), interleukin-8 (IL-8) and tumour necrosis factor-?, assayed on a multiplex platform, and C-reactive protein (CRP), assayed commercially. Relationships were assessed with multiple linear regression analyses.Serum 25(OH)D was positively associated with plasma GSH (? ± SE: 0·002 ± 0·0004) and negatively associated with plasma Eh GSSG (? ± SE: -0·06 ± 0·01) and Cys (? ± SE: -0·01 ± 0·003) (P < 0·001 for all); statistical significance remained after adjusting for age, gender, race, percentage body fat and traditional cardiovascular risk factors (P = 0·01-0·02). The inverse relationship between serum 25(OH)D and CRP was confounded by percentage body fat, and full adjustment for covariates attenuated serum 25(OH)D relationships with other inflammatory markers to nonstatistical significance.Serum 25(OH)D concentrations were independently associated with major plasma thiol/disulphide redox systems, suggesting that vitamin D status may be involved in redox-mediated pathophysiology.

| S-EPMC4115025 | biostudies-literature

Four thiol-oxidoreductases involved in the formation of disulphide bonds in the Streptomyces lividans TK21 secretory proteins.

Project description:BackgroundBacterial secretory proteins often require the formation of disulphide bonds outside the cell to acquire an active conformation. Thiol-disulphide oxidoreductases are enzymes that catalyse the formation of disulphide bonds. The bacterium Streptomyces lividans is a well-known host for the efficient secretion of overproduced homologous and heterologous secretory proteins of industrial application. Therefore, the correct conformation of these extracellular proteins is of great importance when engineering that overproduction.ResultsWe have identified four acting thiol-disulphide oxidoreductases (TDORs) in S. lividans TK21, mutants in all TDOR candidates affect the secretion and activity of the Sec-dependent alpha-amylase, which contains several disulphide bonds, but the effect was more drastic in the case of the Sli-DsbA deficient strain. Thus, the four TDOR are required to obtain active alpha-amylase. Additionally, only mutations in Sli-DsbA and Sli-DsbB affect the secretion and activity of the Tat-dependent agarase, which does not form a disulphide bond, when it is overproduced. This suggests a possible role of the oxidised Sli-DsbA as a chaperone in the production of active agarase.ConclusionsEnzymes involved in the production of the extracellular mature active proteins are not fully characterised yet in Streptomyces lividans. Our results suggest that the role of thiol-disulphide oxidoreductases must be considered when engineering Streptomyces strains for the overproduction of homologous or heterologous secretory proteins of industrial application, irrespective of their secretion route, in order to obtain active, correctly folded proteins.

| S-EPMC6657201 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data