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Hi I am working on a Varian 500MHz NMR. I am attempting to use the pulse sequence satxfer1D to measure the rate constant of an exchanging system. I am not using solvent suppression and I am having trouble understanding the physics behind the "mixing time" after the 90 pulse. I also do not know how many iterations the selective pulse should go through because it seems that if I turn the mixing time off and just use a selective pulse, and then a pw90, I will still obtain the same spectrum no matter how many iterations the selective pulse train goes through. Does anyone have any thoughts/ideas?

asked Sep 29 '11 at 09:54

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njmagarian
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STD NMR is a powerful and fast method used to study protein-ligand interactions, focusing on the signals of the ligand. Besides, values including KD, the association and dissociation kinetics (Koff and Kon respectively) can be obtained from STD NMR study as well.

STD-NMR is a simple and fast method to screen ligand and characterize protein binding and it does not require expensive stable isotope or radioisotope labeling. Even to monitor low receptor concentration at low molecule concentration, STD can be used via non-scalar magnetization transfer, while NMR spectroscopy requires a much higher concentration of proteins that are labeled stable isotopes. In an STD-NMR experiment (as shown in the figure), the di?erence spectrum is obtained by subtracting a spectrum where the protein is selectively saturated (on-resonance) with signal intensities ISAT from that without saturation (off-resonance) with intensities I0. And the difference in intensities (ISTD=I0-ISAT) indicates molecule binding. Creative Biostructure MagHelix™ Saturation Transfer Differences (STD) NMR, hope helpful.

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answered Jun 29 '16 at 19:30

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Justin Frank
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The mixing time is a T1 rho filter to reduce the protein background signal and should have little effect on the STD signal. The filter works well for larger proteins, but may have to be increased for smaller proteins.

We typically use the default settings for the saturation pulse train, see the original Mayer & Meyer article (JACS, 2001, 123, 6108-6117) for more details on this. We also select satmode='ny' and depend only on the DPFGSE sequence for water suppression.

Finally, we have found that the experiment works best in D2O.

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answered Oct 05 '11 at 07:38

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Eugene DeRose
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