Study of Structures and Allosteric Transition
of Lactose Repressor Mutants by X-ray Crystallography
an abstract by Hongli Zhan
Lactose repressor (LacI), a homotetramer of 150 KD, has long been a paradigm
for the allosteric regulation of protein function. Without lactose, tetrameric
LacI binds to its target sites upstream of the Lac operon and inhibits
transcription. A conformation change upon the binding of the inducer
allolactose decreases operator affinity to nonspecific levels. The vacation
of LacI from Lac operon allows the expression of LacZ, Y, and A genes.
Mutagenesis and in vivo screening identify two LacI mutants: L148F and
S151P, both at the core pivot. In vitro characterization indicates that
L148F binds IPTG 4-fold tighter than wild-type LacI (WT), binds O1 40-fold
weaker than WT, and is much less stable to (in?) urea than WT. However,
the allosteric transition is retained in L148F because the operator is
released by IPTG. However, S151P exhibits a higher affinity for O1 and
a lower sensitivity to IPTG. One postulation is that crowding due to
L148F shifts the conformation to a state favorable for inducer binding,
whereas S151P shifts the protein to a state where LacI binds to the operator.
X-ray crystallography is currently being used to determine their structures.
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