Brownian motion, spin diffusion and protein structure determination in solution.

This paper presents my recollections on the development of protein structure determination by NMR in solution from 1968 to 1992. The key to success was to identify NMR-accessible parameters that unambiguously determine the spatial arrangement of polypeptide chains. Inspired by work with cyclopeptides, model considerations showed that enforcing short non-bonding interatomic distances imposes «ring closure conditions» on polypeptide chains. Given that distances are scalar parameters, this indicated an avenue for studies of proteins in solution, i.e., under the regime of stochastic rotational and translational motions at frequencies in the nanosecond range (Brownian motion), where sharp pictures could not be obtained by photography-related methods. Later-on, we used distance geometry calculations with sets of inter-atomic distances derived from protein crystal structures to confirm that measurements of short proton-proton distances could provide atomic-resolution structures of globular proteins. During the years 1976-1984 the following four lines of research then led to protein structure determination by NMR in solution. First, the development of NMR experiments enabling the use of the nuclear Overhauser effect (NOE) for measurements of interatomic distances between pairs of hydrogen atoms in proteins. Second, obtaining sequence-specific resonance assignment solved the "phase problem" for protein structure determination by NMR. Third, generating and programming novel distance geometry algorithms enabled the calculation of atomic-resolution protein structures from limited sets of distance constraints measured by NMR. Fourth, the introduction of two-dimensional NMR provided greatly improved spectral resolution of the complex spectra of proteins as well as efficient delineation of scalar and dipole-dipole H-H connectivities, thus making protein structure determination in solution viable and attractive.