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International Journal of Molecular... Apr 2020Leber's hereditary optic neuropathy (LHON, MIM#535000) is the most common form of inherited optic neuropathies and mitochondrial DNA-related diseases. The pathogenicity...
Leber's hereditary optic neuropathy (LHON, MIM#535000) is the most common form of inherited optic neuropathies and mitochondrial DNA-related diseases. The pathogenicity of mutations in genes encoding components of mitochondrial Complex I is well established, but the underlying pathomechanisms of the disease are still unclear. Hypothesizing that oxidative stress related to Complex I deficiency may increase protein -glutathionylation, we investigated the proteome-wide -glutathionylation profiles in LHON ( 11) and control ( 7) fibroblasts, using the GluICAT platform that we recently developed. Glutathionylation was also studied in healthy fibroblasts ( 6) after experimental Complex I inhibition. The significantly increased reactive oxygen species (ROS) production in the LHON group by Complex I was shown experimentally. Among the 540 proteins which were globally identified as glutathionylated, 79 showed a significantly increased glutathionylation ( < 0.05) in LHON and 94 in Complex I-inhibited fibroblasts. Approximately 42% (33/79) of the altered proteins were shared by the two groups, suggesting that Complex I deficiency was the main cause of increased glutathionylation. Among the 79 affected proteins in LHON fibroblasts, 23% (18/79) were involved in energetic metabolism, 31% (24/79) exhibited catalytic activity, 73% (58/79) showed various non-mitochondrial localizations, and 38% (30/79) affected the cell protein quality control. Integrated proteo-metabolomic analysis using our previous metabolomic study of LHON fibroblasts also revealed similar alterations of protein metabolism and, in particular, of aminoacyl-tRNA synthetases. -glutathionylation is mainly known to be responsible for protein loss of function, and molecular dynamics simulations and 3D structure predictions confirmed such deleterious impacts on adenine nucleotide translocator 2 (ANT2), by weakening its affinity to ATP/ADP. Our study reveals a broad impact throughout the cell of Complex I-related LHON pathogenesis, involving a generalized protein stress response, and provides a therapeutic rationale for targeting -glutathionylation by antioxidative strategies.
Topics: Adenosine Triphosphate; Adult; Aged; Disease Susceptibility; Electron Transport Complex I; Female; Fibroblasts; Humans; Male; Middle Aged; Mitochondria; Models, Molecular; Optic Atrophy, Hereditary, Leber; Protein Conformation; Protein Processing, Post-Translational; Protein S; Proteome; Proteomics; Reactive Oxygen Species; Signal Transduction; Structure-Activity Relationship; Young Adult
PubMed: 32344771
DOI: 10.3390/ijms21083027 -
Biophysical Reviews Dec 2023Lipid-protein interactions are normally classified as either specific or general. Specific interactions refer to lipid binding to specific binding sites within a... (Review)
Review
Lipid-protein interactions are normally classified as either specific or general. Specific interactions refer to lipid binding to specific binding sites within a membrane protein, thereby modulating the protein's thermal stability or kinetics. General interactions refer to indirect effects whereby lipids affect membrane proteins by modulating the membrane's physical properties, e.g., its fluidity, thickness, or dipole potential. It is not widely recognized that there is a third distinct type of lipid-protein interaction. Intrinsically disordered N- or C-termini of membrane proteins can interact directly but nonspecifically with the surrounding membrane. Many peripheral membrane proteins are held to the cytoplasmic surface of the plasma membrane via a cooperative combination of two forces: hydrophobic anchoring and electrostatic attraction. An acyl chain, e.g., myristoyl, added post-translationally to one of the protein's termini inserts itself into the lipid matrix and helps hold peripheral membrane proteins onto the membrane. Electrostatic attraction occurs between positively charged basic amino acid residues (lysine and arginine) on one of the protein's terminal tails and negatively charged phospholipid head groups, such as phosphatidylserine. Phosphorylation of either serine or tyrosine residues on the terminal tails via regulatory protein kinases allows for an electrostatic switch mechanism to control trafficking of the protein. Kinase action reduces the positive charge on the protein's tail, weakening the electrostatic attraction and releasing the protein from the membrane. A similar mechanism regulates many integral membrane proteins, but here only electrostatic interactions are involved, and the electrostatic switch modulates protein activity by altering the stabilities of different protein conformational states.
PubMed: 38192346
DOI: 10.1007/s12551-023-01166-2 -
JACC. Case Reports Aug 2023
PubMed: 37614328
DOI: 10.1016/j.jaccas.2023.101944 -
Transfusion and Apheresis Science :... Aug 2019Although suspected conceptually in the 60 s, Protein C and Protein S activities in hemostasis were investigated and reported from the mid-80 s, followed by the... (Review)
Review
Although suspected conceptually in the 60 s, Protein C and Protein S activities in hemostasis were investigated and reported from the mid-80 s, followed by the discovery of Thrombomodulin, an endothelial cell membrane associated protein, playing the most important heamostatic role. These 3 proteins act in regulating thrombogenesis and protecting against thrombo-embolic events. When blood is activated, any trace of circulating thrombin is captured by Thrombomodulin in the microcirculation, making thrombin become an anticoagulant through its capacity to activate Protein C to Activated Protein C, which operates as a sentinel in blood coagulation, in the form of a complex with free Protein S, to block any new blood activation site, and more especially circulating activated Factors V and VIII. Protein S not only acts as the Activated Protein C cofactor, but also as the cofactor of Tissue Factor Pathway Inhibitor. In addition, it has some functions in the complement pathway through its binding to C4b-BP. Another capability of activated protein C is to lower fibrinolytic activity, as the Activated Protein C Inhibitor is also known as Plasminogen Activator Inhibitor 3. The Protein C-Protein S system becomes less efficient in the presence of mutated Factor V (Factor V-Leiden or other variants), which is resistant to its inactivating effect. Other pathologies linked to this system concern the development of allo- or auto-antibodies to Protein S or to thrombin, which can generate severe thrombotic complications in affected patients. Some antithrombotic drugs have originated from this regulatory system. Protein C or Protein S concentrates are used for treating deficient patients. Activated Protein C (especially in patients with sepsis) or Thrombomodulin are proposed as antithrombotic medications. Most importantly, congenital or acquired Protein C or Protein S deficiencies are associated with severe recurrent thrombotic events. From the clinical standpoint most of the patients are heterozygous, as homozygosity is almost incompatible with life in the absence of a continuous and efficient treatment. Laboratory investigation of this highly complex system involves many different specialized assays for measuring these 3 proteins' activities, their antigenic content or their genetic sequence. The Protein S in-vitro anticoagulant activity is weak and contrasts with its high antithrombotic role in-vivo, showing that diagnostic assays have not yet succeeded in reproducing all the natural mechanisms for evidencing the anticoagulant role of Protein S. This paradoxal notion is discussed and illustrated in this manuscript as well is a revisit of the major characteristics and pathophysiological functions of the Protein C-Protein S-Thrombomodulin system; the associated pathologies; and the main laboratory tools available for clinical diagnosis. In respect to future perspectives, we also focused on developing more significant and relevant assays, especially for Protein S, thanks to the understanding of its biological roles.
Topics: Animals; Blood Coagulation; Humans; Protein C; Protein S; Signal Transduction; Thrombomodulin
PubMed: 31256946
DOI: 10.1016/j.transci.2019.06.008 -
Lipids in Health and Disease Jun 2023Population-based studies investigating the association between blood coagulation markers and non-alcoholic fatty liver disease (NAFLD) are rare. Thus, we aimed to...
BACKGROUND
Population-based studies investigating the association between blood coagulation markers and non-alcoholic fatty liver disease (NAFLD) are rare. Thus, we aimed to investigate the relationship between the Fatty Liver Index (FLI) as a measure of hepatic steatosis and plasma concentrations of antithrombin III, D-dimer, fibrinogen D, protein C, protein S, factor VIII, activated partial thromboplastin time (aPTT), quick value and international thromboplastin time (INR) in the general population.
METHODS
After the exclusion of participants with anticoagulative treatment, 776 participants (420 women and 356 men, aged 54-74 years) of the population-based KORA Fit study with analytic data on hemostatic factors were included in the present analysis. Linear regression models were used to explore the associations between FLI and hemostatic markers, adjusted for sex, age, alcohol consumption, education, smoking status, and physical activity. In a second model, additional adjustments were made for the history of stroke, hypertension, myocardial infarction, serum non-HDL cholesterol levels, and diabetes status. In addition, analyses were stratified by diabetes status.
RESULTS
In the multivariable models (with or without health conditions), significantly positive associations with FLI were obtained for plasma concentrations of D-dimers, factor VIII, fibrinogen D, protein C, protein S, and quick value, while INR and antithrombin III were inversely associated. These associations were weaker in pre-diabetic subjects and largely disappeared in diabetic patients.
CONCLUSION
In this population-based study, an increased FLI is clearly related to changes in the blood coagulation system, possibly increasing the risk of thrombotic events. Due to a generally more pro-coagulative profile of hemostatic factors, such an association is not visible in diabetic subjects.
Topics: Male; Humans; Female; Factor VIII; Antithrombin III; Protein S; Protein C; Blood Coagulation; Hemostatics; Anticoagulants; Fibrinogen
PubMed: 37386502
DOI: 10.1186/s12944-023-01854-8 -
Journal of Thrombosis and Haemostasis :... Apr 2023The complex reactions of blood coagulation are balanced by several natural anticoagulants resulting in tuned hemostasis. During several decades, the knowledge base of... (Review)
Review
The complex reactions of blood coagulation are balanced by several natural anticoagulants resulting in tuned hemostasis. During several decades, the knowledge base of the natural anticoagulants has greatly increased and we have also learned about antiinflammatory and cytoprotective activities expressed by antithrombin and activated protein C (APC). Some coagulation proteins have also been found to function as anticoagulants; e.g., thrombin when bound to thrombomodulin activates protein C. Another example is factor V (FV), which in addition to being a procofactor to FVa has emerged as an anticoagulant. The discovery of APC resistance, caused by FVLeiden, as a thrombosis risk factor resulted in the identification of FV as an APC cofactor working in synergy with protein S in the regulation of FVIIIa in the Xase complex. More recently, a natural anticoagulant FV splice isoform (FV-Short) was discovered when investigating the East Texas bleeding disorder. In FV-Short, the truncated B domain exposes a high-affinity binding site for tissue factor pathway inhibitor alpha (TFPIα), and together with protein S a high-affinity trimolecular complex is generated. The FXa-inhibitory activity of TFPIα is synergistically stimulated by FV-Short and protein S. The circulating FV-Short/protein S/TFPIα complex concentration is normally low (≈0.2 nM) but provides an anticoagulant threshold. In the East Texas bleeding, the concentration of the complex, and thus the threshold, is increased 10-fold, which results in bleeding manifestations. The anticoagulant properties of FV were discovered during investigations of individual patients and follow the great tradition of bed-to-bench and bench-to-bed research in the coagulation field.
Topics: Humans; Anticoagulants; Protein C; Factor V; Protein S; Blood Coagulation
PubMed: 36746318
DOI: 10.1016/j.jtha.2023.01.033 -
Vox Sanguinis Mar 2024Deficiencies of protein C (PC) or protein S (PS) are rare diseases, characterized by mutations in the PC or PS genes, which encode plasma serine proteases with...
BACKGROUND AND OBJECTIVES
Deficiencies of protein C (PC) or protein S (PS) are rare diseases, characterized by mutations in the PC or PS genes, which encode plasma serine proteases with anti-coagulant activity. Severe PC or PS deficiencies manifest in early life as neonatal purpura fulminans, a life-threatening heamorrhagic condition requiring immediate treatment. First-line treatment involves replacement therapy, followed by maintenance with anti-coagulants. Replacement therapy with specific protein concentrates is currently only limited to PC, and therefore, a PC + PS concentrate represents a useful addition to therapeutic options, particularly for severe PS deficiency. Further, the production of a PC + PS concentrate from unused plasma fractionation intermediates would impact favourably on manufacturing costs, and consequently therapy prices for patients and health systems.
MATERIALS AND METHODS
Several chromatographic runs were performed on the same unused plasma fractionation intermediates using different supports to obtain a PC/PS concentrate. The best chromatographic mediums were chosen, in terms of specific activity and recovery. A full process of purification including virus inactivation/removal and lyophilization steps was set up.
RESULTS
The final freeze-dried product had a mean PC concentration of 47.75 IU/mL with 11% of PS, and a mean specific activity of 202.5 IU/mg protein, corresponding to over 12,000-fold purification from plasma.
CONCLUSION
The development of a novel concentrated PC/PS mixture obtained from a waste fraction of other commercial products could be used for its potential therapeutic role in the management of neonatal purpura fulminans pathology.
Topics: Infant, Newborn; Humans; Purpura Fulminans; Protein C Deficiency; Protein C; Protein S; Plasma
PubMed: 38018260
DOI: 10.1111/vox.13567 -
European Journal of Obstetrics,... Apr 2024One serious side effect of combined oral contraceptives (COCs) is venous thromboembolism. Reduced activity in activated protein C-related coagulation pathways is...
OBJECTIVE
One serious side effect of combined oral contraceptives (COCs) is venous thromboembolism. Reduced activity in activated protein C-related coagulation pathways is attributable to low protein S activity in one-third of Japanese patients with deep vein thrombosis. Herer, we quantified the behavior of protein S-specific activity in response to dienogest (DNG) and COCs using the protein S-specific activity assay system to explore its potential utility as a thrombosis marker.
STUDY DESIGN
This was a prospective cohort study. Female patients aged 20 - 49 years who were starting drug treatment for endometriosis using DNG or COCs were enrolled. Blood samples were taken before treatment and at the first, third, and sixth months of treatment. To analyze the primary endpoints, changes in total protein S antigen levels, total protein S activity, and protein S-specific activity from baseline to each time point were estimated using a linear mixed-effects model. All statistical analyses were performed in the SAS software version 9.4 (SAS Institute, Cary, NC). A two-sided P < 0.05 was considered statistically significant.
RESULTS
64 patients took DNG and 34 patients took COCs. Protein S-specific activity did not change significantly from baseline in the six months after treatment started in either group. In the DNG group, total protein S activity and total protein S antigen levels increased slightly from baseline levels after the treatment. The means for total protein S activity and total protein S antigen levels in the COC group remained within reference limits, but they both decreased markedly in the first month and stayed low. Protein S-specific activity in four women remaind below the reference limit throughout the whole study period, suggesting they may have potential protein S deficiencies.
CONCLUSION
The effects of DNG on protein S were negligible, though both total protein S activity and antigen levels decreased soon after COC treatment began and remained low. As there was no VTE event during the study, further studies with larger numbers of patients will be needed to confirm that protein S-specific activity can be a surrogate maker of VTE risk.
Topics: Humans; Female; Contraceptives, Oral, Combined; Endometriosis; Prospective Studies; Nandrolone
PubMed: 38340593
DOI: 10.1016/j.ejogrb.2024.01.028 -
Journal of Medicine and Life Jan 2023Miscarriage in the first and second trimesters of pregnancy is very common, and coagulopathy can be a contributing factor. Protein C and S deficiency are rare, inherited...
Miscarriage in the first and second trimesters of pregnancy is very common, and coagulopathy can be a contributing factor. Protein C and S deficiency are rare, inherited disorders that can increase the risk of thrombophilia. Women with these deficiencies have a higher risk of developing blood clots in the placenta, which can lead to placental insufficiency and, ultimately, to a miscarriage. We aimed to compare the levels of protein C and protein S in pregnant females with recurrent first and second-trimester pregnancy loss and normal pregnant females. We performed a detailed history, examination, and various lab tests on a cohort of 40 females with a history of recurrent first and second-trimester abortions visiting an outpatient clinic at a multi-specialty hospital in Kashmir, India. All the findings were compared with 40 women with normal pregnancies. 10% of the participants had low protein C and S levels (P=0.277), out of whom 75% (p<0.001) had intrauterine growth retardation (IUGR) on ultrasound with 67% (p<0.001) having reduced doppler flow in the umbilical artery. 0.05% of participants had isolated protein S deficiency with no concomitant IUGR seen. Patients with protein C and S deficiencies were treated with heparin and progesterone and followed up for pregnancy outcomes. Screening for protein C and S deficiency is mandatory in all cases of recurrent pregnancy loss. Treatment with low molecular weight heparin and progesterone should be initiated to ensure good fetal outcomes and prevent post-partum/postoperative catastrophic venous thromboembolism events.
Topics: Pregnancy; Female; Humans; Abortion, Habitual; Protein C; Pregnant Women; Progesterone; Embryo Loss; Placenta; Fetal Growth Retardation
PubMed: 36873128
DOI: 10.25122/jml-2022-0267 -
Blood Coagulation & Fibrinolysis : An... Jun 2022The members of a Caucasian family were genetically analyzed on suspicion of hereditary protein S deficiency. A novel mutation, c.1904T>C, associated with severe...
The members of a Caucasian family were genetically analyzed on suspicion of hereditary protein S deficiency. A novel mutation, c.1904T>C, associated with severe quantitative protein S deficiency was found. The novel PROS1 mutation was identified by sequencing of the PROS1 gene coding sequence. The identified c.1904T>C point mutation results in p.Phe635Ser amino acid exchange, which is located in the Laminin G-like 2 domain of protein S. Computational analysis indicates that this amino acid exchange affects the correct folding of the protein S antigen. Furthermore, this mutation is located in a region of the Laminin G-like 2 domain where changes in the amino acid sequence often result in decreased secretion. We postulate that the novel p.Phe635Ser mutation might lead to an incorrect folding, and thus, to a strongly impaired secretion of this protein S variant. We named this novel variant protein.
Topics: Amino Acids; Humans; Laminin; Mutation; Pedigree; Protein S; Protein S Deficiency
PubMed: 34939974
DOI: 10.1097/MBC.0000000000001120