Topology of phage lambda receptor protein. Mapping targets of proteolytic cleavage in relation to binding sites for phage or monoclonal antibodies.
Phage lambda receptor protein of Escherichia coli (LamB protein or maltoporin ) was purified in a mild detergent and subjected to prolonged proteolysis by either trypsin or subtilisin. Cleavage occurred at a limited number of sites without affecting the trimeric structure of the protein. Fragments could be dissociated only by heating in sodium dodecyl sulfate to 100 degrees C. The positions of purified fragments were determined with respect to the uncleaved 421-residue polypeptide by chemical analyses. The regions containing target sites were mapped around residues 159, 203, 245, and 370. Based on kinetics of appearance of the different peptides, early cleavage events occurred at sites near residues 159, 203, and 245 and could be distinguished from late events around residue 370. Information regarding the topological orientation of the cleavage sites could be obtained from the effect of in vitro proteolysis on the ability of the protein to bind phage lambda or monoclonal antibodies. Loss of phage lambda neutralizing activity coincided with early cleavage events, whereas loss of antigenic determinants, known to be exposed at the cell surface, appeared late. Cleavage regions are thus likely to be exposed at the cell surface, a conclusion compatible with the location of mutations affecting the interaction of LamB protein with phage in vivo.
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D010582:Bacteriophage lambda / Q000378:metabolism*; D001665:Binding Sites; D007700:Kinetics; D010446:Peptide Fragments / Q000378:metabolism; D018272:Porins; D011991:Receptors, Virus / Q000378:metabolism*; D013381:Subtilisins / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D010446:Peptide Fragments; D018272:Porins; D011991:Receptors, Virus; C016532:maltoporins; D013381:Subtilisins; D014357:Trypsin