Physicochemical Properties of Crystalline Clostridium Botulinum Type a Toxin*

Interest in the chemical nature and mode of action of bacterial toxins has been renewed by the simultaneously announced crystallization of botulinal (1) and tetanal toxins (2) and the recent purification of several toxoids (3-5). Of these, crystalline Clostridium botulinurn type A toxin alone has been submitted to an apparently complete elementary and amino acid analysis (6), as well as to preliminary physicochemical characterization (7-9) and immunological study (10). This toxin has been found to be a typical protein exhibiting no unique composition or physicochemical properties and is apparently devoid of a prosthetic group. A preliminary report of the molecular weight and homogeneity of botulinal toxin prepared by the method of Lamanna et al. (1) has already been made (9). Further details of the physicochemical characterization of this toxin and of the effect of the method of preparation on the molecular kinetic and serological properties of this substance are given in this report. Investigation of the botulinal toxin-antitoxin reaction has also been undertaken. Data to be published separately indicate that serologically the toxin acts as a single substance (10). The original procedure for the crystallization of botulinal toxin (1) has been modified in several ways, including the omission of a step involving shaking of the crude acid-precipitated toxin with chloroform (11). However, much study had already been made of the properties of crystalline toxin of maximum potency prepared by the procedure first announced. A later method described by Abrams, Kegeles, and Hottle (7) differs from both the above procedures and also omits shaking with chloroform. In view of the suggestion (8) that partial denaturation may result from the use of chloroform in the purification of the toxin, some data on the physicochemical and serological characteristics of toxin prepared both by the original and the modified methods of Lamanna et al. are included in this communication.