Crystallization of genetically engineered active maltose-binding proteins, including an immunogenic viral epitope insertion.
Three mutants of the maltose- or maltodextrin-binding protein encoded by the malE gene of Escherichia coli, with extensive genetic changes, have been purified and crystallized in different crystal forms. Two of these mutant proteins, MalE178 and MalE341, carry net deletions of seven and 13 residues, respectively, near the surface of the molecule. These mutations have very little effect on either the transport activity of the mutant strains or the sugar-binding activity of the purified mutant proteins. The third mutant protein involves the insertion of an 11-residue peptide of the C3 epitope from type 1 poliovirus VP1 protein into the MalE178 deletion mutant, with retention of essentially all the biological properties of the wild-type and the immunological properties of the C3 epitope. We are undertaking three-dimensional structure analysis in order to understand how the protein accommodates these large changes in its surface structure and how the C3 epitope retains its immunological properties in this new environment. The same system could be used to determine easily the structures of other peptide epitopes, especially those in proteins with unknown structures.
Department of Biochemistry, Rice University, Houston, TX 77020.
L E
LE
Rodseth
P
P
Martineau
P
P
Duplay
M
M
Hofnung
F A
FA
Quiocho
D018528:ATP-Binding Cassette Transporters; D000595:Amino Acid Sequence; D001692:Biological Transport; D002213:Capsid / Q000235:genetics / Q000276:immunology*; D036022:Capsid Proteins; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D003460:Crystallization; D000939:Epitopes; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D005818:Genetic Engineering; D008320:Maltose / Q000378:metabolism*; D058619:Maltose-Binding Proteins; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D009154:Mutation; D033902:Periplasmic Binding Proteins; D017955:Poliovirus / Q000235:genetics* / Q000276:immunology; D011134:Polysaccharides / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D014961:X-Ray Diffraction
D018528:ATP-Binding Cassette Transporters; D036022:Capsid Proteins; D002352:Carrier Proteins; D000939:Epitopes; D029968:Escherichia coli Proteins; C461619:MalE protein, E coli; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D033902:Periplasmic Binding Proteins; D011134:Polysaccharides; D011994:Recombinant Proteins; C442540:VP1 protein, Poliovirus; C461599:maltose transport system, E coli; D008320:Maltose; C008315:maltodextrin