Computational protocol: A Metabolically-Stabilized Phosphonate Analog of Lysophosphatidic Acid Attenuates Collagen-Induced Arthritis

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Protocol publication

[…] The inhibition model (competitive, noncompetitive, or mixed type inhibition) was evaluated by graphical analysis with Lineweaver–Burk double reciprocal plots (1/[velocity] versus 1/[substrate]) and Cornish-Bowden plot ([substrate]/[velocity] versus [Inhibitor]). These plots were generated by the measurement of ATX activity in the presence of different BrP-LPA concentrations (0–10 μΜ) and increased amounts of 16:0 LPC (100, 200, 400 μΜ). The inhibition constants (Ki) and IC50 values were determined using the mean values obtained from two independent experiments. The relative remaining ATX activity or LPA levels in the presence of the BrP-LPA was expressed as a percentage of the corresponding control value (ATX activity, LPA levels without inhibitor). The IC50 value was determined graphically by plotting the log concentration of BrP-LPA versus relative remaining ATX activity or LPA levels of each test sample using the sigmoid dose–response fitting method (Prism® software). The IC50 (concentration of BrP-LPA sufficient to inhibit ATX activity or diminish LPA levels by 50%) was calculated by the equation f =  y0+a/(1+exp(-(x-x0)/b)) given by SigmaPlot 11.0 (Systat Software, Inc., San Jose, CA, USA).Ki was calculated using the equation of Cheng and Prusoff: Ki = IC50/1+[S]/Km , where Ki is the binding affinity of the inhibitor, IC50 is the functional strength of the inhibitor, [S] is substrate concentration and Km is the concentration of substrate at which enzyme activity is half maximal. Whereas the IC50 value for a compound may vary between experiments depending on experimental conditions, Ki is an absolute value. Alternatively the Ki values were calculated graphically as follows: The velocity of ATX (v) was determined at two or more substrate concentrations and over a range of inhibitor concentrations (I). In a plot of 1/v against I, data for each substrate concentration fall on straight lines that intersect at I = −Ki and 1/v = 1/Vmax (competitive inhibition) and at I = −Ki and 1/v = [1−(Ki/Ki')]/Vmax (mixed type inhibition). […]

Pipeline specifications

Software tools Dr Fit, SigmaPlot
Applications Drug design, Miscellaneous
Organisms Mus musculus, Homo sapiens
Diseases Angiolymphoid Hyperplasia with Eosinophilia, Arthritis, Arthritis, Experimental, Arthritis, Rheumatoid, Joint Diseases