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PDBsum entry 3e1f
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References listed in PDB file
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Key reference
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Title
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Direct and indirect roles of his-418 in metal binding and in the activity of beta-Galactosidase (e. Coli).
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Authors
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D.H.Juers,
B.Rob,
M.L.Dugdale,
N.Rahimzadeh,
C.Giang,
M.Lee,
B.W.Matthews,
R.E.Huber.
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Ref.
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Protein Sci, 2009,
18,
1281-1292.
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PubMed id
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Abstract
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The active site of ss-galactosidase (E. coli) contains a Mg(2+) ion ligated by
Glu-416, His-418 and Glu-461 plus three water molecules. A Na(+) ion binds
nearby. To better understand the role of the active site Mg(2+) and its ligands,
His-418 was substituted with Asn, Glu and Phe. The Asn-418 and Glu-418 variants
could be crystallized and the structures were shown to be very similar to native
enzyme. The Glu-418 variant showed increased mobility of some residues in the
active site, which explains why the substitutions at the Mg(2+) site also reduce
Na(+) binding affinity. The Phe variant had reduced stability, bound Mg(2+)
weakly and could not be crystallized. All three variants have low catalytic
activity due to large decreases in the degalactosylation rate. Large decreases
in substrate binding affinity were also observed but transition state analogs
bound as well or better than to native. The results indicate that His-418,
together with the Mg(2+), modulate the central role of Glu-461 in binding and as
a general acid/base catalyst in the overall catalytic mechanism. Glucose binding
as an acceptor was also dramatically decreased, indicating that His-418 is very
important for the formation of allolactose (the natural inducer of the lac
operon).
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