Rapid Determination of Fast Protein Dynamics from NMR Chemical Exchange Saturation Transfer Data

Abstract Functional motions of 15 N‐labeled proteins can be monitored by solution NMR spin relaxation experiments over a broad range of timescales. These experiments however typically take of the order of several days to a week per protein. Recently, NMR chemical exchange saturation transfer (CEST) experiments have emerged to probe slow millisecond motions complementing R 1 ρ and CPMG‐type experiments. CEST also simultaneously reports on site‐specific R 1 and R 2 parameters. It is shown here how CEST‐derived R 1 and R 2 relaxation parameters can be measured within a few hours at an accuracy comparable to traditional relaxation experiments. Using a “lean” version of the model‐free approach S 2 order parameters can be determined that match those from the standard model‐free approach applied to 15 N R 1 , R 2 , and { 1 H}‐ 15 N NOE data. The new methodology, which is demonstrated for ubiquitin and arginine kinase (42 kDa), should serve as an effective screening tool of protein dynamics from picosecond‐to‐millisecond timescales.