SOME PROTEIN PROPERTIES IMPORTANT FOR ENZYME CATALYSIS - New Trends in Enzyme Catalysis and Biomimetic Chemical Reactions - page 138

with time constants ranging from 0.1 to several nanoseconds with the total amplitude
between 130 and 340 nm (0.4 - 1.0 kcal/mole).
The first direct evidence of the intramolecular mobility of hemeprotein globules was
obtained by the spin and Mössbauer labeling methods (Frolov et al., 1977; Belonogova
et al., 1978; Park et al., 1981, 1982; Myo et al., 1983; Likhtenshtein and Kotelnikov,
1983). The experiments were carried out on dry and moistened powders, which excluded
any motion by the macromolecule as a whole. The atoms were incorporated into the
heme group in myoglobin and hemoglobin. Given that the rigid heme ring is bound to
the protein by numerous contacts, it is evident that anharmonic motion of heme above
200 K is related to the intramolecular mobility of the protein globule. This mobility
appears only at a critical degree value of hydration samples. The increase of mobility
recorded by NGR-spectroscopy correlates with the data for isotopic H-D whose
relatively higher amplitude must be accompanied by the displacement of the helical
polypeptide chain. Such an unharmonic nanosecond motion with Å also
was recorded at temperature T > 210 K in myoglobin using spin and fluorescence
labeling methods (Likhtenshtein and Kotelnikov, 1983; Likhtenshtein, 1988; and
references therein). The flexibility of the cavity of the myoglobin active site is evidenced
by the mobility of a spin probe, a derivative of isocyanate attached to the heme group in
the single crystal. At room temperature the mobility parameters are that is about

kcal/mole and
NMR relaxation studies can provide detailed information pertaining to the internal
dynamics in proteins on a time scale from milliseconds to picoseconds (Section 1.6.2.).
The and spin-lattice relaxation rate and heteronuclear NOE ' s are

sensitive to high frequency motion while the spin-spin relaxation rate

is a function of much slower processes.
The NMR relaxation technique was used to investigate the backbone dynamics of
staphylococcal nuclease (S. Nase) complexed with a ligand and and labeled
uniformly with (Kay et al., 1989). The relaxation parameters and NOE's were
obtained for over 100 assigned backbones amid nitrogen in the proteins. Information on
internal motions was extracted from experimental data using the model-free approach
(Lipardi and Szabo, 1982). High values of the order parameters characterizing
the extent of rapid bond motion and the correlation time of protein intramolecular
daynamics were determined for and turns and loops. These values as
well as the spin-lattice relaxation rate of did not correlate with the temperature B-
factor calculated from the X-ray analysis. The authors explained this discrepancy by
suggesting different timescales for the different methods. In fact, the B-factor
characterizes not only dynamic processes but also a disordering owing to the presence of
a large number of nearly isoenergetic conformational substrates (Frauenfeldr, 2001;
Frauenfelder et al., 1991). No correlation between rapid small amplitude motions and
secondary structure for S.Nase was found. In contrast, line widths suggest a possible
correlation between secondary structure and motions on the millisecond time scale
monitoring by the measurement spin-spin realaxation rate. The loop region between
residues 42 and 56 appears to be considerably more flexible on the slow time scale than
the rest of the protein. The solid state and solution state NMR studies of the rapid
and highly restricted backbone dynamics of Staphylococcal nuclease indicated the