Pocket Guide to Biomolecular NMR

Michaeleen Doucleff ·Mary Hatcher-Skeers · Nicole J. Crane

Pocket Guide toBiomolecular NMR

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ISBN 978-3-642-16250-3 e-ISBN 978-3-642-16251-0DOI 10.1007/978-3-642-16251-0Springer Heidelberg Dordrecht London New York

© Springer-Verlag Berlin Heidelberg 2011This work is subject to copyright. All rights are reserved, whether the whole or partof the material is concerned, specifically the rights of translation, reprinting, reuse ofillustrations, recitation, broadcasting reproduction on microfilm or in any other way, andstorage in data banks. Duplication of this publication or parts thereof is permitted onlyunder the provisions of the German Copyright Law of September 9, 1965, in its currentversion, and permission for use must always be obtained from Springer. Violations areliable to prosecution under the German Copyright Law.The use of general descriptive names, registered names, trademarks, etc. in thispublication does not imply, even in the absence of a specific statement, that such namesare exempt from the relevant protective laws and regulations and therefore free forgeneral use.

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Springer is part of Springer Science+Business Media (www.springer.com)

Dr. Michaeleen DoucleffNational Institutes of Health (NIH)National Institute of Diabetes &Digestive & Kidney Deseases(NIDDK)

Dr. Mary Hatcher-SkeersThe Claremont CollegesWM Keck Science CenterNorth Mills Avenue 92591711 Claremont CaliforniaUSAmhatcher@jsd.claremont.eduCenter Drive 31

20892-0510 Bethesda Maryland

Buildg.5

USAmcdoucleff@gmail.com

Dr. Nicole J. CraneHenry M. Jackson Foundation for theAdvancement of Military MedicineNaval Medical Research CenterDept. Regenerative MedicineRobert Grant Avenue 50320910 Silver Spring MarylandUSANicole.Crane@med.navy.mil

Preface

. . .energy is applied to the sample in the form of short, intense pulses, andnuclear signals are observed after the pulses are removed. The effects whichresult can be compared to the free vibration or “ringing” of a bell. . .

Eric Hahn, 1953, Physics Today 6, 4–9

The essence of NMR is quite simple. You place a sample in a magneticfield, apply another field, and then collect the electromagnetic wavegenerated by the sample. No need to ionize, crystallize, burn, or evenheat up the sample. A plain-vanilla solution of a compound in water isall you need to identify the substance and characterize many of its keybiophysical properties.

Unfortunately, the elegance and beauty of NMR is often lost whenstudents open a textbook. Even the simplest introductions on NMRimmediately immerse students in mathematics and theories laced withconstants and concepts that lack tangible, macroscopic manifestations.For example, many textbooks begin by defining the energy of a nucleusin a magnetic field, E = −γ�mBo, a simple equation that is clearand understandable to all. However, take a closer look at this equality.Hidden inside the constants and variables are sophisticated conceptsthat are tough for many of us to relate to. Take “γ ” for instance.Wikipedia defines “γ ” (i.e., the gyromagnetic ratio) as the ratio of thenucleus’s magnetic dipole moment to its angular momentum. Ridingthe swings at a carnival definitely helps us to relate to angular momen-tum. But what exactly is a magnetic dipole moment, and why would

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taking the ratio of the magnetic dipole moment and the angular momentbe proportional energy? Very quickly, the theory and mathematicsbehind NMR plunges into ideas and phenomenon that aren’t presentin our macroscopic world, making them difficult to grasp, remember,and use.

In contrast to the labyrinth of equations in NMR textbooks, perform-ing actual NMR experiments is remarkably easy. You place the samplein the magnet, set a couple of parameters, and hit “go.” After a fewcommands at the keyboard, your spectrum appears on the screen witha peak for each hydrogen in the compound. Can’t get much easier thanthat! So why can’t the theory of NMR be that simple too?

That, indeed, is the purpose of this book—to bring the commonsense and practicality of experimental NMR to the theory of NMR.To accomplish this task, we set three major goals:

1. Preserve the simplicity of NMR: Instead of using equations andcomplex formulism to explain why nuclei behave as they do in amagnetic field, we simply describe how nuclei act during NMRexperiments. We minimize jargon and keep the descriptions short,pithy, and easy to understand. Students with minimal experiencein chemistry and biology should easily follow the entire book. Theonly equations used in the main text are the sine and exponentialfunctions.

2. Add a hefty dose of intuition to NMR: In place of mathematics andformulism, we use concrete analogies to which readers can relate,making concepts easy to assimilate and use. In this regard, we havecreated a new framework for explaining NMR experiments. Insteadof trying to define “spin,” we simply state that nuclei “ring” in mag-netic fields like tiny bells. This bell analogy, which is employedthroughout the book, has never been used to explain NMR andmakes it surprisingly easy to learn advanced NMR concepts, suchas dipole–dipole coupling and relaxation theory.

3. Flatten the learning curve: The “bell” analogy also provides a newlanguage for discussing NMR experiments. Because this languageis based on an intuitive model of NMR, students quickly master

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it. In other words, this new approach flattens the steep learningcurve of NMR and makes NMR accessible to students at all levels,even those with little experience in spectroscopy, quantum mechan-ics, or physics. Furthermore, the intuitive perspective presented inthis book will help advanced students remember and integrate moremathematical explanations of NMR into their experimental designsand analyses.

The small size and fast pace of the book makes it well suited as acompanion to traditional biophysics or biochemistry textbooks at theundergraduate or graduate levels. However, we hope that the book willalso be useful for professional researchers in the biological scienceswho are interested in collaborating with NMR spectroscopists or ingarnering a better understanding of research articles that present NMRdata.

Finally, one note about the style and tone of the book. The bookis written to be read. Therefore, we use a light, fun style that willhopefully hold your attention from the initial descriptions of one-dimensional experiments to the final pages covering CPMG sequences.We hope that you enjoy it!

Michaeleen DoucleffMary Hatcher-Skeers

Nicole J. Crane

Contents

1 Atomic Bells and Frequency Finders . . . . . . . . . . . 11.1 Chemical Choirs . . . . . . . . . . . . . . . . . . . . 11.2 Essentials of Electromagnetism . . . . . . . . . . . . 31.3 Electromagnetic Microsensors . . . . . . . . . . . . . 51.4 Frequency Finders . . . . . . . . . . . . . . . . . . . 81.5 Basics of One-Dimensional NMR . . . . . . . . . . . 16Further Reading . . . . . . . . . . . . . . . . . . . . . . . 18

2 Bonded Bells and Two-Dimensional Spectra . . . . . . . 192.1 Introduction to Coupling . . . . . . . . . . . . . . . . 192.2 Bonded Bells: J-Coupling . . . . . . . . . . . . . . . 222.3 NMR Maps: Two-Dimensional Spectra . . . . . . . . 272.4 The 1H–15N HSQC: Our Bread and Butter . . . . . . 342.5 Hidden Notes: Creating Two-Dimensional Spectra . . 37References and Further Reading . . . . . . . . . . . . . . . 42

3 Neighboring Bells and Structure Bundles . . . . . . . . 433.1 Bumping Bells: Dipole-Dipole Coupling . . . . . . . 443.2 Atomic Meter Stick: The NOE . . . . . . . . . . . . 513.3 Into “Three-D” . . . . . . . . . . . . . . . . . . . . . 573.4 Adult “Connect-the-Dots:” HNCA . . . . . . . . . . 623.5 Putting the Pieces Together: A Quick Review . . . . . 66

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3.6 Wet Noodles and Proteins Bundles: Buildinga Three-Dimensional Structure . . . . . . . . . . . . 69

References and Further Reading . . . . . . . . . . . . . . . 73

4 Silencing of the Bells: Relaxation Theory Part One . . . 754.1 Nothing Rings Forever: Two Paths to Relax . . . . . . 764.2 Relaxation: Ticket to the Protein Prom . . . . . . . . 814.3 Oh-My, How Your Field Fluctuates . . . . . . . . . . 844.4 Blowing Off Steam and Returning to Equilibrium: T1 924.5 Loosing Lock-Step: Coherence and T2 . . . . . . . . 97References and Further Reading . . . . . . . . . . . . . . . 105

5 Relaxation Theory Part Two: Moving Atomsand Changing Notes . . . . . . . . . . . . . . . . . . . . 1075.1 Keeping the Terms Straight . . . . . . . . . . . . . . 1085.2 NMR Dynamics in a Nutshell: The Rules of Exchange 1115.3 Two States, One Peak: Atoms in the Fast

Lane of Exchange . . . . . . . . . . . . . . . . . . . 1135.4 Two States, Two Peaks: Atoms in the Slow

Lane of Exchange . . . . . . . . . . . . . . . . . . . 1165.5 Two States, One Strange Peak: Atoms

in Intermediate Exchange . . . . . . . . . . . . . . . 1175.6 Tumbling Together: Rotational Correlation Time (τc) 1235.7 Summary . . . . . . . . . . . . . . . . . . . . . . . . 128Further Reading . . . . . . . . . . . . . . . . . . . . . . . 129

6 Protein Dynamics . . . . . . . . . . . . . . . . . . . . . . 1316.1 Dynamics Analysis by NMR: Multi-channel

Metronomes, Not a GPS . . . . . . . . . . . . . . . . 1326.2 Elegant Simplicity: Lipari and Szabo Throw

Out the Models . . . . . . . . . . . . . . . . . . . . . 1336.3 Wagging Tails and Wiggling Bottoms: Local

Versus Global Motion . . . . . . . . . . . . . . . . . 1356.4 Measuring Fast Motion: Model Free Analysis . . . . 1376.5 Changing Directions on the Track: Refocusing Pulses 146

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6.6 Measuring Intermediate Motion: CPMGRelaxation Dispersion Analysis . . . . . . . . . . . . 148

6.7 Measuring Slow Motion: Z-Exchange Spectroscopy . 1516.8 Measuring Motion Summary . . . . . . . . . . . . . 154References and Further Reading . . . . . . . . . . . . . . . 155

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157