Pl. Zhang et al., UNUSUAL LACK OF INTERNAL MOBILITY AND FAST OVERALL TUMBLING IN OXIDIZED FLAVODOXIN FROM ANACYSTIS-NIDULANS, Journal of Molecular Biology, 272(3), 1997, pp. 443-455
Anacystis nidulans flavodoxin, an electron-transfer protein containing
a flavin mononucleotide (FMN) molecule as its prosthetic group, has a
redox potential for the oxidized/semiquinone equilibrium close to tha
t of free flavin. Whereas the redox potential for the semiquinone/hydr
oquinone equilibrium is more negative. To gain an understanding of the
contribution of mobility to redox potential modulation, we studied th
e backbone mobility of the oxidized A, nidulans flavodoxin at pH 6.6,
303 K by N-15 NMR relaxation measurements. The spin-lattice relaxation
rate constants (R-N(N-z)=1/T-1), spin-spin relaxation rate constants
(R-N(N-x,N-y)=1/T-2) and H-1-N-15 nuclear Overhauser effects (NOE) wer
e obtained for 143 of the 166 protonated backbone N-15 nuclei and for
the FMN N3 nucleus without ambiguity. The N-15 T-1, T-2 and NOE data w
ere analyzed by reduced spectral density mapping, and the so-called mo
del-free approach. In contrast to most other proteins studied with N-1
5 relaxation experiments, we found an almost complete absence of inter
nal mobility. The overall correlation time of this 169-residue flavodo
xin (>19 kDa) is significantly shorter (7.4 to 7.8 ns) than that of ot
her proteins of this size, suggesting that the absence of internal mob
ility is correlated with faster overall rotational diffusion. The unif
ormity of the motional parameters along the backbone is in strong cont
rast to the crystallographic B-factors, which vary significantly along
the sequence in this and other flavodoxins. The NMR relaxation parame
ters are primarily sensitive to rotational diffusive motions of the N-
H bond vectors, while the crystallographic B-factors would be sensitiv
e to translational internal motions as well. However, the large B-fact
ors in this protein may originate from crystal packing and crystal lat
tice disorder. The relatively fast overall tumbling results in sharp N
MR resonances. Hence, much larger proteins with such favorable dynamic
behavior could be excellent candidates for studies by NMR. (C) 1997 A
cademic Press Limited.