Abstract
The maltose-binding protein (MBP) of Escherichia coli is the periplasmic receptor of the maltose transport system. Previous studies have identified amino acid substitutions in an alpha/beta loop of the structure of MBP that are critical for the in vivo folding. To probe genetically the structural role of this surface loop, we generated a library in which the corresponding codons 32 and 33 of malE were mutagenized. The maltose phenotype, which correlates with a biologically active structure of MBP in the periplasm, indicated a considerable variability in the loop residues compatible with a correct in vivo folding pathway of the protein. By the same genetic screens, we characterized loop-variant MBPs associated with a defective periplasmic folding pathway and aggregated into inclusion bodies. Heat-shock induction with production of misfolded loop variants was examined using both lon-lacZ and htrA-lacZ fusions. We found that the extent of formation of inclusion bodies in the periplasm of E. coli, from misfolded loop variant MBPs, correlated with the level of heat-shock response regulated by the alternate heat-shock sigma factor, sigma 24.
MeSH terms
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ATP-Binding Cassette Transporters*
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ATP-Dependent Proteases
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Amino Acid Sequence
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Bacterial Proteins*
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Calcium-Binding Proteins*
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Carrier Proteins / chemistry*
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Carrier Proteins / genetics
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Databases, Factual
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Escherichia coli
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Escherichia coli Proteins*
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Heat-Shock Proteins / metabolism
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Hot Temperature
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Inclusion Bodies / chemistry*
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Inclusion Bodies / ultrastructure
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Lac Operon
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Maltose-Binding Proteins
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Molecular Sequence Data
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Monosaccharide Transport Proteins*
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Mutagenesis, Site-Directed
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Periplasmic Binding Proteins*
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Periplasmic Proteins*
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Protease La*
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Protein Structure, Secondary
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Protein Structure, Tertiary
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Serine Endopeptidases / metabolism
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Structure-Activity Relationship
Substances
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ATP-Binding Cassette Transporters
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AraF protein, E coli
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Bacterial Proteins
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Calcium-Binding Proteins
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Carrier Proteins
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Escherichia coli Proteins
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Heat-Shock Proteins
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LivK protein, E coli
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MalE protein, E coli
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Maltose-Binding Proteins
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Monosaccharide Transport Proteins
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Periplasmic Binding Proteins
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Periplasmic Proteins
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RbsB protein, E coli
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galactose-binding protein
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histidine-binding protein
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lysyl-arginyl-ornithine-binding protein, bacteria
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maltose transport system, E coli
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sbp protein, E coli
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sulfate-binding protein, bacteria
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ATP-Dependent Proteases
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DegP protease
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Serine Endopeptidases
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Lon protein, E coli
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Protease La