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This viral RNA was extensively studied and undergoes rigorous conformational exchange on the fast NMR chemical shift time scale.
Previous studies have shown that the S- and D-states of IscU give rise to different spectral signatures on the slow chemical shift time scale.
The NMR experiments, because the exchange process is fast on the chemical shift time scale, do not provide information on the structures of the sparsely populated states.
The observation that many resonances in the free BaDHPS were unobservable suggested that BaDHPS undergoes motions on the intermediate chemical shift time scale (i.e., μs ms).
Stepwise titrations of SL3ESS3 into N-labeled UP1 reveal slow exchange on the NMR chemical shift time scale, consistent with the nanomolar binding affinity determined by calorimetry (Table 1).
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However, the general tendency is that as the lifetime of a complex is increased (i.e. binding becomes relatively stronger), the exchange regime on the chemical-shift time scale goes from fast to intermediate to slow.
In the case of lactose binding to gal-1, we observed that gal-1 resonances were merely chemically shifted during the titration with lactose, which is what one would expect with fast exchange on the chemical-shift time scale.
In our case, gal-1 resonances initially may be shifted somewhat by interaction with GRG, but are primarily broadened due to the exchange process which falls in the intermediate exchange regime on the NMR chemical-shift time scale [ 22].
We do this in a way that is similar to HSQC chemical-shift mapping [ 23], which is performed when binding interactions occur in the fast or slow exchange regimes on the chemical-shift time scale.
The difference in Kd values is consistent with the significant resonance broadening we observed with gal-1 binding to GRG, placing the interaction in the intermediate exchange regime on the chemical-shift time scale [ 22].
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com