NMR Experiments for Studies of Dilute and Condensed Protein Phases: Application to the Phase-Separating Protein CAPRIN1.

Intrinsically disordered proteins (IDPs) or regions of intrinsic disorder in otherwise folded proteins (IDRs) play important roles in many different biological processes, including formation of biological condensates via liquid-liquid phase separation. NMR spectroscopy is a powerful tool for obtaining site-specific structural and dynamical information on IDPs/IDRs, and recent efforts have focused on the development of experiments for atomic-resolution studies of these molecules. These include triple-resonance experiments that are based on CO-direct detection of magnetization, exploiting increased sensitivity of cryogenically cooled probes.

Hyperinsulinism-Causing Mutations Cause Multiple Molecular Defects in SUR1 NBD1.

The sulfonylurea receptor 1 (SUR1) protein forms the regulatory subunit in ATP sensitive K (K) channels in the pancreas. SUR proteins are members of the ATP binding cassette (ABC) superfamily of proteins. Binding and hydrolysis of MgATP at the SUR nucleotide binding domains (NBDs) lead to channel opening.

Measurement of signs of chemical shift differences between ground and excited protein states: a comparison between H(S/M)QC and R1rho methods.

Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion NMR spectroscopy has emerged as a powerful tool for quantifying the kinetics and thermodynamics of millisecond exchange processes between a major, populated ground state and one or more minor, low populated and often invisible 'excited' conformers. Analysis of CPMG data-sets also provides the magnitudes of the chemical shift difference(s) between exchanging states (|Deltavarpi|), that inform on the structural properties of the excited state(s). The sign of Deltavarpi is, however, not available from CPMG data.

Structural studies of FF domains of the transcription factor CA150 provide insights into the organization of FF domain tandem arrays.

FF domains are poorly understood protein interaction modules that are present within eukaryotic transcription factors, such as CA150 (TCERG-1). The CA150 FF domains have been shown to mediate interactions with the phosphorylated C-terminal domain of RNA polymerase II (phosphoCTD) and a multitude of transcription factors and RNA processing proteins, and may therefore have a central role in organizing transcription. FF domains occur in tandem arrays of up to six domains, although it is not known whether they adopt higher-order structures.

Measuring the signs of 1H(alpha) chemical shift differences between ground and excited protein states by off-resonance spin-lock R(1rho) NMR spectroscopy.

Analysis of Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion NMR profiles provides the kinetics and thermodynamics of millisecond-time-scale exchange processes involving the interconversion of populated ground and invisible excited states. In addition, the absolute values of chemical shift differences between NMR probes in the exchanging states, |Delta omega|, are also extracted. Herein, we present a simple experiment for obtaining the sign of (1)H(alpha) Delta omega values by measuring off-resonance (1)H(alpha) decay rates, R(1rho), using weak proton spin-lock fields.