BioPHYSICAL chemistry

A second approach is to use solid-state NMR (McDermott 2004). The

protein is no longer in solution and so does not tumble, and a broad

distribution would be seen due to the anisotropy of the chemical shift.

However, the contribution can be eliminated with spinning of the sample

at the magic angle of 54.74° (Figure 16.12). At this angle, the dipole –

dipole dependence of 1–3 cos^2 θgives an average value of zero and so there

is no contribution to the spectrum. The application of this technique requires

that the rotation frequency be not less than the width of the spectrum.

This requires the use of kilohertz rotation frequencies for the sample in

the magnet, which are achieved with the use of gas-driven spinners.

Together, the development of these techniques for NMR provides the

opportunity to address a major question in proteomics, namely how to

356 PART 2 QUANTUM MECHANICS AND SPECTROSCOPY

0.2

26

25

27

1.2 1.0 0.8 0.6 0.4

13 C (ppm)

(^1) H (ppm)
(c)
14
12
10
16
1.0 0.8 0.6 0.4
13 C (ppm)
(^1) H (ppm)
(a) (b)
1.2 1.0 0.8 0.6 0.4
(^1) H (ppm)
(d) (e)
1.2 1.0 0.8 0.6 0.4
(^1) H (ppm)
L192 δ 1
L148 δ 1
L233 δ 1
L201 δ 1
L192 δ 2
L207 δ (^2) L136 δ 2
L148 δ 2
L184 δ 1
L81 δ L47 δ^1 L178 δ 1
L47 δ 1
L38 δ 1
L81 δ
L48 δ 1
L58 δ
L197 δ 2
L233 δ 2
L192 δ 1
L207 δ 1
L148 δ 1
L106 δ 1
L197 δ 2
L176 δ 1
L148 δ 2
L136 δ 2
L192 δ (^2) L184 δ 1
L136 δ 1
L136 δ 2
L112 δ 2
L192 δ 1
L207 δ 1
I157 δ 1
I109 δ 1
I233 δ (^1) I141 δ 1
I165 δ 1
I215 δ 1
I49 δ (^1) I212 δ 1 I137 δ 1
I59 δ 1 I67 δ^1
I94 δ 1
670-kDa α 7 β 7 β 7 α 7 20S proteasome
670-kDa α 7 β 7 β 7 α 7 20S proteasome CP
360-kDa α 7 α 7 double ring
360-kDa α 7 α 7 double ring
21-kDa αmonomer
670-kDa α 7 β 7 β 7 α 7 CP 21-kDa αmonomer
I64 δ 1
I70 δ 1
L197 δ 2
L106 δ 1
L148 δ 1
L176 δ 1
L207 δ 2
L201 δ 1 L148 δ^2
L47 δ 1
L88 δ 1
L38 δ 1
L48 δ 1
L69 δ 1
unassigned
L233 δ 1
L192 δ 2 L207 δ 2 L112 δ^2
L184 δ 1
L201 δ 1
L48 δ 1
L47 δ 1
L69 δ 1
L58 δ L38 δ 1
L207 δ 1
Figure 16.11Structure of the proteasome and TROSY spectra. (a) The structure of the proteasome
shows a overall barrel shape formed from rings of two different subunits. (b–e) The methyl-based
TROSY spectra, which are color-coded according to the assigned region, show very sharp peaks
that allow assignments despite the large size of the complex. From Sprangers and Key (2007).