I am trying to run a STD-NMR to determine the position of my drug binding with a protein called COX-2. How much of protein should I use? The protein comes at 5000 unit.
Hi Song,
It depends on many factors, besides of course the MW of the protein. For example, the experiment is highly sensitive to the kinetics of the binding of your drug. I would recommend you to start with no less than 1 mg in 500-600 uL in the NMR tube, and something around 500uM-2mM of your drug.
I'm not entirely sure of what you mean by the "position" of your drug. If you want to determine "binding mode" that would be feasible by STD NMR. If you want to detect the "location" in the protein surface that is another story. In that case you could use docking calculations using the X-ray structure of COX-2, either to predict the location, or to predict the bound conformation and binding mode, if the site of your drug is known. Then you could "validate" your 3D model of the complex by running CORCEMA-ST calculations. Additionally, you can run DEEP-STD NMR experiments (multifrecuency STD NMR protocol), to figure out which type of amino acids are contacting your ligand in the bound state.
Best of luck!
Hi Song,
It depends on many factors, besides of course the MW of the protein. For example, the experiment is highly sensitive to the kinetics of the binding of your drug. I would recommend you to start with no less than 1 mg in 500-600 uL in the NMR tube, and something around 500uM-2mM of your drug.
I'm not entirely sure of what you mean by the "position" of your drug. If you want to determine "binding mode" that would be feasible by STD NMR. If you want to detect the "location" in the protein surface that is another story. In that case you could use docking calculations using the X-ray structure of COX-2, either to predict the location, or to predict the bound conformation and binding mode, if the site of your drug is known. Then you could "validate" your 3D model of the complex by running CORCEMA-ST calculations. Additionally, you can run DEEP-STD NMR experiments (multifrecuency STD NMR protocol), to figure out which type of amino acids are contacting your ligand in the bound state.
Best of luck!
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