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Lecture Seven: Structures from NMR
[Based on Chapter 3 – Berg,
Tymoczko & Stryer]
(Figures in red are for the 7th Edition)
 Nuclear Magnetic Resonance (NMR) can reveal the
______________________ of macromolecules
 NMR is unique in giving the structure in _____________
 Background
 Table 3-4, page 98 (Table 3-4, page 103)
 Many nuclei of atoms have an intrinsic magnetism
 The most significant for biological systems are:
 1H
13
C
15
N
31
P
 These isotopes can be used in NMR to study
macromolecular structures
 1H is useful for _____________
 Extensively found in biological systems

31
P is useful for _____________
 Remember - (NOT found in proteins)
 The magnetism in these nuclei comes from one property:
the SPIN of their protons
 Example: 1H has a spin of _____
 Figure 3-48, page 98 (3-44, page 103)
 The spin of 1H generates a magnetic moment
 In an applied magnetic field this adopts one of two
orientations
   orientated with the field
   orientated against the field
 The applied magnetic field strength is Bo
 Very powerful magnets are needed
 E is the ___________________ between  and 
 E is proportional to Bo
 E is in the radio frequency range
 1H nuclei in the  state can be excited into the  state
 This requires a _____________ of radio-frequency
energy

o =
 Ho
2
 o is the radio-frequency - Important
  is the magnetogyric ratio for a given nucleus
 ___________ around 1H alter the magnetism that the nucleus
experiences
 Creates a local chemical environment
 Examples:
 A methyl group (CH3) is one chemical
environment
 An aromatic ring (C6H6) would be a different
chemical environment
 Electrons shield protons from the applied field
 Ho =
Bo ( 1   )
 Ho is the _________ magnetic field strength
  is the shielding factor
 1H nuclei in different chemical environments will have
different o values
 Differences in o values are very small
 Scaled as a term, 
 Called the ________________
 Measured as parts per million (ppm)
 Figure 3-49, page 99 (3-45, page 104)
 1H nuclei in different chemical environments have specific
regions of chemical shift
 Examples:
 Methyl groups:

 =
0  1.5 ppm
 Aromatic groups:

 =
6  7.5 ppm
 The local magnetic field is further altered
 ____________________ bonds to neighbouring
nuclei with magnetic moments
 Nuclei affect each other
 This is called _________________
 Requirements for observing spin-spin coupling
 Neighbouring nuclei must be in different chemical
environments
 Note: True even for two seemingly identical groups
 Nuclei must usually be _____________ bonds apart
 Otherwise the effect is too small to be seen
 Observing the effect
 Nuclei excited to the  state must lose energy to return to
the  state
 This is called _____________
 Occurs via an interaction with other near
neighbour magnetic nuclei
 An NMR spectrum (Fig. 3-49) is the observation of
 state protons falling back to the  state
 Two forms of relaxation occur:
 Relaxation through bonds
 Relaxation through space
 Relaxation can also be observed by 2-D NMR
Spectroscopy
 2-D NMR Spectroscopy
 General Information
 Spectra are drawn as a 2-D ‘contour’ map of peaks
 Peaks on the ___________ are the 1-D NMR spectra
 Peaks off the diagonal show where relaxation
interactions have occurred (see above)
 2-D Correlation Spectroscopy (COSY)
 Provides information on the through bond connections
between residues
 Identifies the _____________
 2-D Nuclear Overhauser Enhancement Spectroscopy
 Usually called NOESY
 Reveals information about through space relaxation
interactions
 Uses Nuclear Overhauser Effect (NOE) data
 This interaction is from nuclei separated in space by < 5Å
 Figure 3-50, page 100 (3-46, page 104)
 The peaks on the diagonal of a NOESY spectrum
are a normal 1-D spectrum
 Used as markers to locate the off-diagonal peaks
 1H nuclei close in space to each other will '________' an
off-diagonal peak
 Off-diagonal peaks are symmetric in appearance
 Off-diagonal peaks show where relaxation has occurred
between 1H nuclei through space
 Figure 3-51, page 100 (3-47, page 105)
 A large number of off-diagonal peaks are present in a
2-D spectrum
 All represent nuclei in close proximity to each other
 Figure 3-52, page 101 (3-48, page 105)
 For atomic structure determination all the NOESY data
must be satisfied
 All 1H nuclei shown by NOESY to be close together
must be close together in the structure
 Together with additional data obtainable from NMR a SET
of solution conformation of a protein can be obtained
 An ________________ of structures
 Figure 3-53, page 101 (3-49, page 105)
 Potential Problems
 No SINGLE STRUCTURE exists for a protein
 Because NOESY data are for a range of lengths
 Methyl groups often dealt with as a SINGLE atom
 Called ________________
 Summary of Lecture Seven:
 Nuclear Magnetic Resonance (NMR) can provide
atomic structures of proteins
 NMR uses nuclei with intrinsic magnetism
 1H is very important for biological studies
 It has a spin of 1/2
 In a large applied magnetic field 1H nuclei adopt one of
two orientations
  with the field
 against the field
 Radio frequencies are used to excite 1H nuclei from  to 
 1H nuclei are in different chemical environments because
of different amounts of surrounding electrons
 Electrons shield nuclei from the applied magnetic field
 A local field results
 The different environments mean 1H nuclei are excited at
different frequencies
 Differences are very small
 Recorded as  ppm (parts per million)
 Neighbouring bonded magnetic nuclei affect each other
 Called spin-spin coupling
 Relaxation is the loss of energy from an excited
nucleus  to 
 2-D NMR can be used to record relaxation data
 Two forms:
 Through bond  COSY
 Through space  NOESY
 COSY shows the peak positions for the amino acid groups
 NOESY shows nuclei that are close in space
 This information is used to obtain atomic structures of
proteins