Enzyme IIA
Glc, encoded
by the
crr gene of the phosphoenolpyruvate:sugar phosphotransferasesystem, plays an important role in regulating intermediary meta
bolism in
Escherichia coli ("cata
boliterepression"). One function involves inhi
bition of induci
ble transport systems ("inducer exclusion"), andwith lactose permease, a galactoside is required for unphosphorylated IIA
Glc binding to cytoplasmic loopsIV/V and VI/VII [Sondej, M., Sun, J. et al. (1999)
Proc. Natl. Acad. Sci. U.S.A. 96, 3525-3530]. Withinside-out mem
brane vesicles containing the permease, [
125I]IIA
Glc binding promoted
by meli
biose exhi
bitsan affinity (
KDIIA) of
![](/images/entities/ap.gif)
1
![](/images/entities/mgr.gif)
M and a stoichiometry of one mole of IIA
Glc per six moles of lactose permease.Both the quantity of [
125I]IIA
Glc bound and the sugar concentration required for half-maximal IIA
Glc binding(
K0.5IIAsug) was measured for eight permease su
bstrates. Differences in maximal IIA
Glc binding are o
bserved,and the
K0.5IIAsug does not correlate with the affinity of LacY for sugar. Furthermore,
K0.5IIAsug does notcorrelate with sugar affinities for various permease mutants. IIA
Glc does not
bind to a mutant (Cys154
![](/images/entities/rarr.gif)
Gly), which is locked in an outwardly facing conformation,
binds with increased stoichiometry to mutantLys131
![](/images/entities/rarr.gif)
Cys, and
binds only weakly to two other mutants which appear to
be predominantly in eitheran outwardly or an inwardly facing conformation. When the latter two mutations are com
bined, sugar-dependent IIA
Glc binding returns to near wild-type levels. The findings suggest that
binding of varioussu
bstrates to lactose permease results in a collection of unique conformations, each of which presents aspecific surface toward the inner face of the mem
brane that can interact to varying degrees with IIA
Glc.