E. Steensma et Cpm. Vanmierlo, STRUCTURAL CHARACTERIZATION OF APOFLAVODOXIN SHOWS THAT THE LOCATION OF THE STABLE NUCLEUS DIFFERS AMONG PROTEINS WITH A FLAVODOXIN-LIKE TOPOLOGY, Journal of Molecular Biology, 282(3), 1998, pp. 653-666
The structural characteristics of Azotobacter vinelandii apoflavodoxin
II have been determined using multidimensional NMR spectroscopy. Apof
lavodoxin has a stable, well-ordered core but its flavin binding regio
n is flexible. The local stability of apoflavodoxin was probed using h
ydrogen/deuterium exchange measurements. The existence of an apoflavod
oxin equilibrium folding intermediate is inferred from the noncoincide
nce of CD and fluorescence unfolding curves obtained for the guanidini
um hydrochloride induced unfolding of apoflavodoxin. We suggest that t
he structured part of the putative intermediate is composed of the ele
ments of secondary structure which have the slowest exchanging amide p
rotons in the native protein. These elements are strands beta 1, beta
3, beta 4 and beta 5 alpha and helices alpha 4 and alpha 5. We propose
that it is a general feature of flavodoxins that the stable nucleus r
esides in the C-terminal part of these proteins. The results on flavod
oxin are compared with those on two sequentially unrelated proteins sh
aring the flavodoxin-like fold: Che Y and cutinase. It is shown that t
he stable nucleus is found in different parts of the flavodoxin-like t
opology. (C) 1998 Academic Press.