Lipoprotein from the outer membrane of Escherichia coli: purification, paracrystallization, and some properties of its free form.
In the envelope of Escherichia coli, is a lipoprotein of molecular weight 7,200 as a major envelope protein. This lipoprotein was previously shown to exist in two different forms in the outer membrane of E. coli: the free form and the boundform, which is covalently linked to the peptidoglycau. The free form of the lipoprotein has been purified and paracrystallized by adding acetone to a sodium dodecyl sulfate solution in the presence of magnesium ion. The paracrystals were needle shaped. An electron micrograph of the negatively stained paracrystals showed a highly ordered ultrastructure. The chemical structure of the free form was compared with that of the bound form by (i) the amino acid composition, (ii) the fatty acid composition, and (iii) the peptide analysis after cyanogen bromide cleavage. The alpha-helical content of the free form of the lipoprotein was measured from the circular dichroism spectrum of the lipoprotein in 0.01% sodium dodecyl sulfate and found to be 87%. Using the purified lipoprotein as antigen, antiserum against the free form of the lipoprotein was obtained. Immunoprecipitation of the lipoprotein with the antiserum was found to be very specific, since only the free form of the lipoprotein was found as a major peak when the antiserum was reacted with the whole envelope proteins solubilized in 0.2% sodium dodecyl sulfate, and the immunoprecipitate thus formed was analyzed by polyacrylamide gel electrophoresis.
D000596:Amino Acids / Q000032:analysis; D001426:Bacterial Proteins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification; D002473:Cell Wall / Q000032:analysis / Q000648:ultrastructure; D005591:Chemical Fractionation; D003460:Crystallization; D000939:Epitopes; D004926:Escherichia coli / Q000648:ultrastructure*; D005227:Fatty Acids / Q000032:analysis; D008074:Lipoproteins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification; D008970:Molecular Weight; D010455:Peptides / Q000032:analysis
D000596:Amino Acids; D001426:Bacterial Proteins; D000939:Epitopes; D005227:Fatty Acids; D008074:Lipoproteins; D010455:Peptides