title: Protein-bound choline is released from the pneumococcal autolytic enzyme during adsorption of the enzyme to cell wall particles.
abstract: The inactive precursor form of the pneumococcal autolytic enzyme cloned in Escherichia coli was isolated by affinity chromatography on Sepharose-linked choline. The enzyme was recovered in an electrophoretically pure and activated form by elution from the affinity column with radioactive choline solution. When radioactive choline was used for elutions, the enzyme protein isolated contained protein-bound choline, at approximately 1 mol of choline per mol of enzyme protein, indicating the presence of a single choline recognition site. Radioactive choline remained bound to the enzyme protein during dialysis, precipitation by trichloroacetic acid or ammonium sulfate, and during gel filtration, but not during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Incubation of the choline-labeled autolysin with pneumococcal cell walls at 0 degrees C resulted in the adsorption of the enzyme to the wall particles and a simultaneous release of free choline from the enzyme protein. It is suggested that the choline molecules that became bound to the enzyme protein during the activation of autolysin are expelled from the choline-binding site and replaced by choline residues from the wall teichoic acid as the autolysin molecules adsorb to their insoluble substrate before the onset of enzymatic wall hydrolysis.
authors.affiliation: Rockefeller University, New York, New York 10021-6399.
authors.forename: Z
authors.initials: Z
authors.lastname: Markiewicz
authors.forename: A
authors.initials: A
authors.lastname: Tomasz
mesh_terms: D000327:Adsorption; D000581:Amidohydrolases / Q000378:metabolism*; D002473:Cell Wall / Q000378:metabolism*; D002794:Choline / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D007700:Kinetics; D008954:Models, Biological; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000378:metabolism*; D013296:Streptococcus pneumoniae / Q000201:enzymology* / Q000235:genetics
chemical_list: D000581:Amidohydrolases; D009238:N-Acetylmuramoyl-L-alanine Amidase; D002794:Choline