Abstract
Lactose permease of Escherichia coli (LacY) with a single-Cys residue in place of A122 (helix IV) transports galactopyranosides and is specifically inactivated by methanethiosulfonyl-galactopyranosides (MTS-gal), which behave as unique suicide substrates. In order to study the mechanism of inactivation more precisely, we solved the structure of single-Cys122 LacY in complex with covalently bound MTS-gal. This structure exhibits an inward-facing conformation similar to that observed previously with a slight narrowing of the cytoplasmic cavity. MTS-gal is bound covalently, forming a disulfide bond with C122 and positioned between R144 and W151. E269, a residue essential for binding, coordinates the C-4 hydroxyl of the galactopyranoside moiety. The location of the sugar is in accord with many biochemical studies.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Amino Acid Substitution
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Binding Sites / genetics
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Biological Transport
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Crystallization
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Cysteine / chemistry
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Cysteine / genetics
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Cysteine / metabolism
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Escherichia coli / genetics
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Escherichia coli / metabolism
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Escherichia coli Proteins / chemistry*
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Escherichia coli Proteins / genetics
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Escherichia coli Proteins / metabolism
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Galactose / chemistry*
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Galactose / metabolism
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Lactose / chemistry
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Lactose / metabolism
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Membrane Transport Proteins / chemistry*
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Membrane Transport Proteins / genetics
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Membrane Transport Proteins / metabolism
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Mesylates / chemistry
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Mesylates / metabolism
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Models, Molecular
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Monosaccharide Transport Proteins / chemistry
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Monosaccharide Transport Proteins / genetics
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Monosaccharide Transport Proteins / metabolism
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Protein Binding
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Protein Conformation
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Protein Structure, Tertiary*
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Substrate Specificity
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Symporters / chemistry
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Symporters / genetics
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Symporters / metabolism
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X-Ray Diffraction
Substances
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Escherichia coli Proteins
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Membrane Transport Proteins
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Mesylates
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Monosaccharide Transport Proteins
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Symporters
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methanethiosulfonate
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lactose permease
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Lactose
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Cysteine
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Galactose