Learning modern protein NMR

Protein NMR Spectroscopy. Principles and Practice by John Cavanagh, Wayne Fairbrother, Arthur G Palmer III and Nicholas J Skelton, Academic Press, 1996, 587 pages, $59.95 hardback (ISBN 0-12-164490-1).NMR has been around for just over 50 years and the first applications to proteins date back more than 30 years. However, it has been only a dozen years since the first structures of very small proteins were solved by NMR. The early phase of this NMR work is covered, for example, in the 1986 text ‘NMR of proteins and Nucleic Acids’ by K Wuthrich. However, since then an enormous number of methodological advances not only have extended the size limit of proteins that can be studied by NMR, but have also increased the attainable level of detail on local structure and internal dynamics. Whereas in the 1980s, structures were derived on the basis of simple two-dimensional (2D) nuclear Overhauser effect spectroscopy (NOESY) and correlation spectroscopy (COSY) spectra, the study of proteins beyond the 10 kDa barrier typically requires the use of a battery of 3D and 4D NMR spectra, carried out on isotopically enriched proteins.Protein NMR Spectroscopy. Principles and Practice is aimed at the spectroscopist who wants to ‘get his/her hands dirty’, not at the structural biologist who wants to figure out what's behind these ‘fuzzy NMR structures’. All four authors have first-hand experience with developing and using the newest NMR methods, dating back to the days when they were working jointly at the Institute of Molecular Biology at Scripps Clinic, La Jolla, California. Their book describes clearly the principles underlying these modern NMR experiments and the intricate mechanisms of a substantial subset of them.The book starts with an introductory chapter, describing magnetic resonance in classical terms and explaining chemical shifts, chemical shift anisotropy, scalar couplings, and the limitations of the classical approach. The next chapter deals with a theoretical description of NMR and lays much of the basis for what follows. Besides going into details regarding the quantum mechanical description and the ever-important density matrix, the chapter provides a very clear description of the product operator formalism which has served as the primary vehicle for the development of many of today's pulse schemes.Before the book dives into the actual multidimensional NMR experiments, it provides two chapters dedicated to a mixture of important concepts, ranging from data acquisition and processing, solvent signal suppression techniques, spin decoupling, and pulsed field gradients to details of how one actually designs a sequence to select the signals of interest. Along with all the theoretical details, this material is interspersed with numerous useful practical hints, and many simple equations and derivations which make it easier to obtain a quantitative grasp.The main interest of Palmer, the principal author of the text, is in the use of NMR to characterise internal motions from relation measurements. It is therefore no surprise that the section dealing with relaxation and dynamics is particularly lucid. All relaxation mechanisms applicable for diamagnetic proteins are discussed in detail, both for the case of laboratory and rotating frame measurements. Cross-relaxation between proximate protons, which constitutes the main source of structural information in protein structure determination, is also described in a manner which starting graduate students may find particularly useful.Although conventional 2D 1H NMR of proteins is no longer particularly fashionable, there are numerous small proteins for which structure determination can be carried out easily using this ‘old’ approach. The authors demonstrate the experiments for human ubiquitin, a very small and well-behaved protein of 76 residues, and all spectra shown in the book have been specifically recorded for this text using the ubiquitin sample. This has the advantage that the reader can immediately evaluate the relative quality and information content of the spectra generated by the wide range of experimental techniques.Whereas there are over a dozen useful homonuclear experiments of proteins, the number of 3D and 4D experiments applicable to isotopically enriched proteins is at least an order of magnitude larger. Although it clearly is not possible to provide a detailed description of all of these techniques, the authors have done an excellent job at picking some of the most representative ones and discussing these in some detail. Important practical details, unique to 3D and 4D NMR, such as the use of aliasing to increase the spectral resolution, are discussed clearly and illustrated in the application of the various schemes to uniformly (13C/15N)-enriched ubiquitin. Although not all of the pulse schemes discussed have incorporated recent advances in spectrometer hardware, pulsed field gradients in particular, the results shown are of high quality and give any spectroscopist new to the field a yard stick of what to expect or, at least, what to shoot for.How does one go from NMR spectra to protein structure? There is no single answer to this question and a detailed discussion of this topic would fill another volume. Cavanagh et al. are therefore necessarily brief on this topic, but nevertheless give an interesting overview of this type of work.Protein NMR is a rapidly moving field and methodological advances continue to outpace the rate at which books can be printed. Nevertheless, the principles are not expected to change much in the foreseeable future, and Protein NMR Spectroscopy. Principles and Practice therefore is strongly recommended to graduate students, post-does, and all those interested in the intricate details of biomolecular NMR spectroscopy. Although the title of the book refers to proteins, the methodology used for studying nucleic acids or other molecules, for that matter, largely overlaps with the material covered and the book should therefore also be of use to NMR spectroscopists outside the protein field.With the enormous amount of detail contained in this book, including nearly 900 equations, it is not surprising that a few errors have slipped through the proof-reading stage. The authors have set up a web page (http://convex.hhmi.columbia.edu/palmer/protein_nmr_spectroscopy) to remedy the problem. The book contains a reasonably complete bibliography, and a superb subject index. Along with detailed explanations of the mechanisms underlying the modern NMR experiments, the book also brings some ‘order’ to the bewildering array of novel NMR pulse sequences. To practitioners of biomolecular NMR, it will prove invaluable for years to come.