Maleimide scavenging enhances determination of protein S-palmitoylation state in acyl-exchange methods

Protein C, a vitamin K-dependent anticoagulant serine protease, is involved in blood coagulation. Activated protein C inactivates Va and VIIIa in purified protein systems and stimulates fibrinolysis by indirectly increasing the level of circulating plasminogen activator. In this process, protein S serve as an important factor for activated protein C. In recent years, excess protein S drives cancer cell proliferation and cell survival through oncogenic receptor Axl (Anexelekto). We determined changes of plasma protein C antigen by using rocket immunoassay both in 50 healthy individuals and 103 distinct hospitalized patients. In healthy individuals protein C antigen(PC:Ag) ranges o.6439- 1.4752 µ/ml. The results showed that plasma protein C antigen was considerably high in 22 diabetes mellitus. In contrast, the PC:Ag was significantly decreased in 19 liver cirrhosis(p< 0.001) and in closely line with serum albumin levels(p< 0.05). In 31 acute leukemias, on the average, there was slightly lower values in PC:Ag, and accompanied with the distribution of significant decrease of PC:Ag values in 5 FAB M5 subtype and in 9 hyperleukocytic leukemias. However, the 3 acute promyelocytic leukemia (APL) with overt laboratory criteria of disseminated intravascular coagulation (DIC) had protein C concentration no lower than the remaining 2 patients with infectious DIC, which suggested the coagulopathy in APL might be due to mechanisms different from other forms of DIC.

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Monoclonal Antibody-Based Enzyme-Linked Immunosorbent Assays (ELISA) for the Measurement of Vitamin K-Dependent Protein S: The Effect of Antibody Immunoreactivity on Plasma Protein S Antigen Determinations

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648514 ◽

1992 ◽

Vol 67 (06) ◽

pp. 631-638 ◽

Cited By ~ 10

Author(s):

Silvana Vigano D’Angelo ◽

Sandra Tombesi ◽

Santica Marcovina ◽

Alberto Albertini ◽

Patrizia Della Valle ◽

...

Keyword(s):

Protein S ◽

Plasma Samples ◽

Potential Influence ◽

Polyclonal Igg ◽

Dependent Protein ◽

Capture Antibody ◽

Antigen Determination ◽

Sample Dilution ◽

Selection Of

SummaryTwo monoclonal antibodies (Mabs) specifically directed to human protein S (PS) - named 5E9E9 and 3B10.25 - were produced and their properties compared to those of 2 previously characterized anti-PS-Mabs (HPS-2 and S10). 3B10.25, similar to S10, was directed to the calcium-free conformation of PS and had virtually identical affinity for free and C4b-binding protein (C4b-BP)-bound PS; 5E9E9 similar to HPS-2, had no calcium-dependency and was selectively directed to free PS. All Mabs were equally reactive to freshly purified and thrombin-cleaved PS. To evaluate the influence of C4b-BP bound PS on PS antigen determinations, ELISA systems employing the four Mabs individually as capture antibody (Ab) and peroxidase-conjugated polyclonal anti-PS IgG as detecting Ab were developed and compared to immunoelectrophoresis (EIA) and to an ELISA employing polyclonal anti-PS IgG as capture and detecting Ab, in the determination of PS in purified systems and in plasma. With all the ELISAs there was parallelism of dilution cuiyes obtained with normal plasma and purified PS; however, supplementation of plasma with purified C4b-BP resulted in loss of parallelism when employing the Mabs directed to free PS as capture Ab. Influence of high C4b-BP on PS antigen determinations was confirmed in a series of plasma samples from patients with C4b-BP levels ranging from 70% to over 200%. Compared to the values obtained with the S10- or 3B10.25 - based ELISAs - which were similar despite a 10-fold difference in sample dilution -plasma PS was underestimated by the ELISAs employing 5E9E9 or HPS-2 while it was overestimated by EIA. In addition, plasma PS and C4b-BP levels were significantly correlated only when it was measured by EIA or the ELISAs employing S10, 3B10.25 or polyclonal IgG. These results highlight the potential influence of high C4b-BP on plasma PS antigen determination. Accurate measurement of PS by ELISA requires selection of antibodies with identical affinity for all plasma PS forms.

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Determination of Protein S-Acylation State by Enhanced Acyl-Switch Methods

Methods in Molecular Biology - Protein Lipidation ◽

10.1007/978-1-4939-9532-5_1 ◽

2019 ◽

pp. 3-11

Author(s):

Charlotte H. Hurst ◽

Dionne Turnbull ◽

Piers A. Hemsley

Keyword(s):

Protein S

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An Impedimetric Approach for COVID-19 Detection

The Analyst ◽

10.1039/d1an01718g ◽

2021 ◽

Author(s):

Yudum Tepeli ◽

Burak Ekrem Citil ◽

U. Anik

Keyword(s):

Protein S ◽

Spike Protein ◽

S Protein ◽

Infection Mechanism ◽

Biosensor System ◽

Electrochemical Approach

In this study, an electrochemical approach for the determination of coronavirus disease (COVID-19) was developed. The biosensor system relied on the spike protein (S-protein) based infection mechanism of the virus...

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First experience with a new assay for activity determination of protein S

Fibrinolysis and Proteolysis ◽

10.1016/0268-9499(94)90631-9 ◽

1994 ◽

Vol 8 ◽

pp. 124 ◽

Cited By ~ 1

Author(s):

C. Wagner ◽

A. Girolami ◽

P. Simeoni ◽

F. Dati

Keyword(s):

Protein S ◽

Activity Determination

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Metal-responsive elements in Pleurotus ostreatus laccase gene promoters

Microbiology ◽

10.1099/mic.0.26360-0 ◽

2003 ◽

Vol 149 (8) ◽

pp. 2155-2162 ◽

Cited By ~ 55

Author(s):

Vincenza Faraco ◽

Paola Giardina ◽

Giovanni Sannia

Keyword(s):

Pleurotus Ostreatus ◽

Copper Ions ◽

Protein S ◽

Start Codon ◽

Gene Promoters ◽

Laccase Gene ◽

Promoter Regions ◽

Molecular Masses ◽

Laccase Genes

Fungal laccase gene transcription is strongly induced by copper ions; notably, some laccase promoters contain multiple putative metal-responsive elements (MREs). Previously, it has been demonstrated that the Pleurotus ostreatus laccase genes poxc and poxa1b are transcriptionally induced by copper, and several putative MREs were found in the promoter regions of these genes, which extend for about 400 nt upstream of the start codon (ATG). Identification of MRE sequences, which are protected by protein binding in the poxc and poxa1b promoter regions, has been achieved by footprinting analyses. Electromobility shift assays led to the evaluation of the ability of the identified MREs to bind protein(s), and the role of specific nucleotides of these elements in complex formation has also been analysed. The formation of complexes between analysed MREs and fungal proteins requires the absence of metal ions. Proteins extracted from fungus grown in copper-depleted medium are able to form complexes with MREs, whilst proteins extracted from fungus grown in copper-containing medium are able to form complexes only in the presence of a metal chelator. Moreover, copper-depleted proteins are unable to form complexes when copper or zinc ions are added. UV-cross-linking analyses led to the determination of the molecular masses of the MRE-binding proteins. In the poxa1b promoter, a GC-rich region, homologous to the core binding site for transcription factor Sp1, decreases the binding affinity of the adjacent MRE, affecting its interactions with fungal protein factors.

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