PMID:9232651

Citation	Sun, J, Frillingos, S and Kaback, HR (1997) Binding of monoclonal antibody 4B1 to homologs of the lactose permease of Escherichia coli. Protein Sci. 6:1503-10
Abstract	The conformationally sensitive epitope for monoclonal antibody (mAb) 4B1, which uncouples lactose from H+ translocation in the lactose permease of Escherichia coli, is localized in the periplasmic loop between helices VII and VIII (loop VII/VIII) on one face of a short helical segment (Sun J, et al., 1996, Biochemistry 35;990-998). Comparison of sequences in the region corresponding to loop VII/VIII in members of Cluster 5 of the Major Facilitator Superfamily (MFS), which includes five homologous oligosaccharide/H+ symporters, reveals interesting variations. 4B1 binds to the Citrobacter freundii lactose permease or E. coli raffinose permease with resultant inhibition of transport activity. Because E. coli raffinose permease contains a Pro residue at position 254 rather than Gly, it is unlikely that the mAb recognizes the peptide backbone at this position. Consistently, E. coli lactose permease with Pro in place of Gly254 also binds 4B1. In contrast, 4B1 binding is not observed with either Klebsiella pneumoniae lactose permease or E. coli sucrose permease. When the epitope is transferred from E. coli lactose permease (residues 245-259) to the sucrose permease, the modified protein binds 4B1, but the mAb has no significant effect on sucrose transport. The studies provide further evidence that the 4B1 epitope is restricted to loop VII/VIII, and that 4B1 binding induces a highly specific conformational change that uncouples substrate and H+ translocation.
Links	PubMed PMC2143751 Online version:10.1002/pro.5560060714
Keywords	Amino Acid Sequence; Antibodies, Bacterial; Antibodies, Monoclonal; Binding Sites; Biological Transport; Epitopes; Escherichia coli/enzymology; Escherichia coli/genetics; Escherichia coli Proteins; Membrane Transport Modulators; Membrane Transport Proteins/antagonists & inhibitors; Membrane Transport Proteins/genetics; Membrane Transport Proteins/immunology; Membrane Transport Proteins/metabolism; Membranes/metabolism; Molecular Sequence Data; Monosaccharide Transport Proteins; Mutagenesis, Site-Directed; Protein Structure, Secondary; Species Specificity; Spheroplasts/metabolism; Symporters
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
CITFR:Q5ETW8		GO:0016021: integral to membrane	ECO:0000247:	UniProtKB:P02920	C		Integral to membrane based on sequence alignment. See Table 1.	complete CACAO 3822
CITFR:Q5ETW8		GO:0005351: sugar:proton symporter activity	ECO:0000247:	UniProtKB:P02920	F		Sugar:hydrogen symporter activity based on sequence alignment. See Table 1.	complete CACAO 3820
CITFR:Q5ETW8		GO:0034219: carbohydrate transmembrane transport	ECO:0000314:		P		Figure 3A shows C. freundii lac permease protein transporting (active transport) melibiose sugar over time compared to E. Coli raffinose permease. The direct assay experiment shows that a conserved sequence of residues is maintained and exhibit similar function between the two species' transmembrane sugar transporters.	complete CACAO 4773
CITFR:Q5ETW8		GO:0015155: lactose transmembrane transporter activity	ECO:0000314:		F		Figure 2 shows interactions involving lac permease.	complete
CITFR:Q5ETW8	enables	GO:0005351: carbohydrate:proton symporter activity	ECO:0000247: sequence alignment evidence used in manual assertion	UniProtKB:P02920	F		Seeded From UniProt	complete
CITFR:Q5ETW8	part_of	GO:0016021: integral component of membrane	ECO:0000247: sequence alignment evidence used in manual assertion	UniProtKB:P02920	C		Seeded From UniProt	complete

References

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