Titration of Drug Using GPC Spin ColumnESIMS Coelution of Multiple Ligands Noncovalently Bound to Protein Target

A titration study was undertaken to determine the extent of non-covalent binding between the inhibitor DBQ (MW 489 Da) and the protein CMVP A144L. This experiment was conducted by incubating a fixed amount (50 pmol) of the protein CMVP A144L with from 0-40 mol excesses of DBQ. The incubated materials were then subjected to GPC spin column analysis, whereby the eluate containing the tightly bound inhibitor was analyzed by ESI-MS under denaturing conditions to quantitate the mole ratio of inhibitor to protease (D/P). Figure 2.25 illustrates the [M+H]1+ region of the ESI mass spectrum for these titration experiments. (Note the characteristic 1:2:1 dibromo isotopic intensity distribution.) Table 2.5 summarizes the quantitative results for these titration experiments by tabulating the [drug]/[protein] (D/P) molar ratios during the incubation reaction and after elution from the spin column. Figure 2.26 graphically illustrates a plot of the D/P mole ratios of the reaction vs the bound mixtures. Note that at low reaction D/P mole ratios, from 0 to 1, the slope of the curve rises rapidly and continuously levels off from reaction D/P mole ratios from 1 to 40. These results suggest that there may be specific non-covalent binding at low D/P mole ratios up to about 0.5 D/P mole ratios, but as the reaction D/P ratio increases less specific non-covalent

Healthy Chemistry For Optimal Health

Healthy Chemistry For Optimal Health

Thousands Have Used Chemicals To Improve Their Medical Condition. This Book Is one Of The Most Valuable Resources In The World When It Comes To Chemicals. Not All Chemicals Are Harmful For Your Body – Find Out Those That Helps To Maintain Your Health.

Get My Free Ebook

Post a comment