Immunochemical Studies with the β-Galactosidase and Structurally Related Proteins of <i>Escherichia Coli</i>

Abstract

Summary Rabbit antisera against purified Escherichia coli β-galactosidase (Gz) have been prepared. These antisera precipitate the enzyme but do not inactivate it. The fact that the enzyme titrates with this antiserum to the same endpoint whether it is determined with lactose or orthonitrophenol β-galactoside as substrate, demonstrates that both activities are associated with a single antigenic species. Inactive extracts (from cells which have not been induced to produce enzyme) as well as active extracts (from cells which possess the enzyme) contain a protein (Pz), inactive as a β-galactosidase which cross-reacts strongly with the antibody to Gz. Pz, although immunologically closely related to Gz is distinguishable by the following properties. Pz will precipitate 80–90% of the antibody directed against Gz, but leave a fraction which will react only with Gz to the exclusion of Pz. In mixtures of Pz and Gz, Gz precipitates preferentially and Pz remains in solution as long as the precipitation of Gz is not complete. Pz is nonantigenic for rabbits whereas Gz is an exceptionally efficient antigen. The following summarizes some other properties of Pz and Gz. Pz and Gz have very similar solubility properties but are separable by the fact that Gz and Pz have appreciable differences in their temperature coefficients of solubility. Gz is more stable than Pz towards the action of heat, ultraviolet irradiation and proteolytic digestion. Gz can be converted into an enzymatically inactive, immunologically identical protein by the action of dilute formalin under proper conditions. These results show that the induced formation (enzymatic adaptation) of β-galactosidase involves the appearance of a new antigen Gz and, therefore, the biosynthesis of a new protein. The significance of the immunological behavior of Pz and Gz is discussed with special reference to problems of protein homogeneity and interpretation of the reactions of complex systems.

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Affiliated Institutions

• Institut Pasteur FR

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