Figure 4 - available via license: CC BY-NC-ND
Content may be subject to copyright.
Effect of QPS and Q3R on clotting times. A) APTT and B) PT were measured using human plasma expressed as ratio of clotting time (sec), as described in materials and methods section. Values are represented as mean± SEM. P values of less than 0.05 were considered as significant with *P<0.05. Data are representative of at least three independent experiments (n=5 in each group).
Source publication
Aims
Design and synthesis of a novel 3, 3', 4', 5, 7-O- pentasulfated Quercetin (QPS) as activator of protein disulfide isomerase (PDI).
Main Methods
Based on an in silico analysis we show that QPS binds to a and b domain of PDI (-7.4 kcal/mol) unlike PDI inhibitor quercetin 3-rutinoside (Q3R) that binds at substrate binding domain b (-8.0 kcal/mo...
Contexts in source publication
Context 1
... 1 H-NMR spectral analysis of proton chemical shifts of QPS indicated downfield shifts for aromatic protons H-2', H-5', H-6, H-8 in the range 8.07-7.39 ppm whereas the signals for OH protons were only observed in quercetin in the range 12.49-9.31 ppm, thereby further establishing the substitution of OH group by sulfate moieties in case of QPS (Fig. ...
Context 2
... [14]. QPS effect on the coagulation rates using APTT (extrinsic) and PT (intrinsic) coagulation pathways showed that addition of exogenous recombinant PDI and QPS independently decreased the coagulation time indicating an increase in the rate of coagulation, however this increase in rate was slightly reversed on addition of PDI and QPS together ( Fig. 4A and 4B). Q3R showed a significant increase in coagulation time indicating J o u r n a l P r e -p r o o f a decrease in rate of coagulation in both APTT and PT assays, while the addition of PDI and Q3R together only marginally increases the rates of coagulation. Prothrombotic affect of QPS indicates its modulation of coagulation cascade ...
Context 3
... insulin turbidity assay to check for PDI activity confirmed that QPS increases the activity (Fig. 3), while Q3R reduces its activity. Coagulation assays done in isolation shows addition of exogenous PDI or QPS to be thrombotic, however incubating both in combination decreases the coagulation rates slightly but it is still less than control ( Figure 4A and 4B). This marginal decrease in rate of coagulation is probably due exogenous QPS activated PDI influencing other coagulation targets. ...
Citations
Extracellular protein disulfide isomerase (PDI) is a promising target for thrombotic-related diseases. Four potent PDI inhibitors with unprecedented chemical architectures, piericones A-D (1-4), were isolated from Pieris japonica. Their structures were elucidated by spectroscopic data analysis, chemical methods, quantum 13C nuclear magnetic resonance DP4+ and electronic circular dichroism calculations, and single-crystal X-ray diffraction analysis. Piericones A (1) and B (2) were nanomolar noncompetitive PDI inhibitors possessing an unprecedented 3,6,10,15-tetraoxatetracyclo[7.6.0.04,9.01,12]pentadecane motif with nine contiguous stereogenic centers. Their biosynthetic pathways were proposed to include a key intermolecular aldol reaction and an intramolecular 1,2-migration reaction. Piericone A (1) significantly inhibited in vitro platelet aggregation and fibrin formation and in vivo thrombus formation via the inhibition of extracellular PDI without increasing the bleeding risk. The molecular docking and dynamics simulation of 1 and 2 provided a novel structure basis to develop PDI inhibitors as potent antithrombotics.
Thromboembolic diseases are a major cause of mortality in human and the currently available anticoagulants are associated with various drawbacks, therefore the search for anticoagulants that have better safety profile is highly desirable. Compounds that are part of the dietary routine can be modified to possibly increase their anticoagulant potential. We show mannose 2,3,4,5,6-O-pentasulfate (MPS) as a synthetically modified form of mannose that has appreciable anticoagulation properties. An in silico study identified that mannose in sulfated form can bind effectively to the heparin-binding site of antithrombin (ATIII) and heparin cofactor II (HCII). Mannose was sulfated using a simple sulfation strategy-involving triethylamine-sulfur trioxide adduct. HCII and ATIII were purified from human plasma and the binding analysis using fluorometer and isothermal calorimetry showed that MPS binds at a unique site. A thrombin inhibition analysis using the chromogenic substrate showed that MPS partially enhances the activity of HCII. Further an assessment of in vitro blood coagulation assays using human plasma showed that the activated partial thromboplastin time (APTT) and prothrombin time (PT) were prolonged in the presence of MPS. A molecular dynamics simulation analysis of the HCII-MPS complex showed fluctuations in a N-terminal loop and the cofactor binding site of HCII. The results indicate that MPS is a promising lead due to its effect on the in vitro coagulation rate.
Communicated by Ramaswamy H. Sarma
