Abstract
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 1H–1H connectivities, thus making protein structure determination in solution viable and attractive.
Highlights
Background from high school and universitiesMy education, leading to a Ph.D. degree, was pursued in Switzerland, which is my home country
In the early days of NMR spectroscopy with biological macromolecules, from 1957 into the 1980s, only a few proteins were accessible for structural studies, either by X-ray crystallography or by NMR in solution, mainly because milligram quantities of the proteins had to be isolated and purified from natural sources
truncated-driven NOE (TOE) measurements correspond to the standard experiment used for small molecules [28], except that the preirradiation had to be highly selective for work with crowded protein 1H NMR spectra; applying the continuous wave (CW) irradiation for variable short time periods was used to record NOE build-up curves and monitor the unwanted contributions from spin diffusion [32]
Summary
‘‘för hans utveckling av kärnmagnetisk resonansspektroskopi för bestämming av biologiska makromolekylers tredimensionella struktur i lösning” (official translation: ‘‘for his development of nuclear magnetic resonance spectroscopy for determining the three-dimensional structure of biological macromolecules in solution”); this is the citation in the Nobel Prize diploma that was handed to me on December 10, 2002. In the early days of NMR spectroscopy with biological macromolecules, from 1957 into the 1980s, only a few proteins were accessible for structural studies, either by X-ray crystallography or by NMR in solution, mainly because milligram quantities of the proteins had to be isolated and purified from natural sources. Protein crystallography was years ahead of NMR for structural studies of biological macromolecules, and we benefited in key issues from their experience, in particular from the early efforts to refine [9] and classify [10] protein crystal structures
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