COMPARATIVE STUDY OF BACTERIAL EXTRACTS BY ANALYTICAL ELECTROPHORESIS1

Sidney D. Rodenberg
1958 Journal of Bacteriology  
Electrophoretic analysis of the complex solutions extractable from microbial cells was first reported by Melnick and Stern (1940) . However, ultracentrifugation has been used much more extensively for the analysis and fractionation of bacterial extracts. Using analytical ultracentrifugation, Schachman et al. (1952) showed the essential similarity of the extracts of a number of species of bacteria. The macromolecules of an extract were found to have sedimentation properties which caused them to
more » ... ove at three different rates in centrifugal fields. Thus, three principal groups of molecules or components were characteristic of all bacterial extracts studied and the three different sedimentation rates of the components were likewise similar in all of the extracts. The validity and usefulness of their findings is evident from the subsequent work of a number of investigators (reviewed by Alexander, 1956) . Electrophoretic examinations of microbial extracts have been largely in support of fractionation studies and the results reported have been more complex than those from ultracentrifugal studies. With extracts of Saccharomyces cerevisiae, Melnick and Stern (1940) found that more components were detectable by analytical electrophoresis than by ultracentrifugation of the same extract. Siegel et al. (1952) described electrophoretic differences between bacteriophage-infected and normal cells of Escherichia coli. Hess and Slade (1955) found the electrophoretic method sufficiently descriptive to permit characterization of a number of closely related streptococci. Although ultracentrifugal patterns of bacterial extracts were not altered by differences in extraction procedures, Hess and Slade (1956) reported electrophoretic differences related to 1 Aided by a grant (G-722) from the National Science Foundation. 2 A portion of the work reported was undertaken while the writer was the recipient of the Orville Paul Phillips fellowship at the University of Pennsylvania. 84 the length of time used for sonic extraction, and Melnick and Stern (1940) noted similar results when different solvents were employed for the extraction. More recently, Hess et al. (1957) found considerable quantitative differences in the composition of three strains of Streptococcus pyogenes. The selection of Proteus vulgaris for intensive electrophoretic study following a survey of a number of bacterial extracts was described by Rodenberg (1957) . The medium in which cells were grown, the temperature of incubation, the degree of aeration during incubation, and the age of the culture were found to affect the electrophoretic pattern of sonic extracts of bacteria. This paper, the first of a series, reports the results of electrophoretic analyses of bacterial extracts representing a majority of the families of the Eubacteriales. These studies were undertaken to evaluate the usefulness of this method for investigations of protein synthesis in microorganisms. After this work was complete and the manuscript in an advanced stage of preparation, the paper of Wagman et al. (1958) appeared. In part, the results reported here are in agreement with their findings. MATERIALS AND METHODS Cultivation of organisms and preparations of extracts. The identities of the organisms which were maintained in the stock culture collection of this laboratory were confirmed shortly before they were used. With the exception of Mycobacterium smegmatis, which was grown in nutrient broth (Difco) containing 1 per cent glucose and 0.05 per cent "Tween 80", all organisms were grown in a medium containing glucose, 0.1 per cent; yeast extract, 1 per cent; tryptone, 1 per cent; and K2HPO4, 0.2 per cent. The buffer used throughout these studies was 0.1 ionic strength phosphate buffer prepared at pH 7 (Miller and Golder, 1950) . Anaerobic organisms were grown in 6 L round bottom flasks containing 5.5 L of medium. Other organisms were grown in 1 L on May 4, 2020 by guest
doi:10.1128/jb.76.1.84-93.1958 fatcat:z3c5taeesveyvb5kaikgc4i6d4