Choline-containing Teichoic Acid As a Structural Component of Pneumococcal Cell Wall and Its Role in Sensitivity to Lysis by an Autolytic Enzyme

Abstract

The cell wall of Diplococcus pneumoniae was isolated and purified and its macromolecular components were characterized in two ways: (a) by selective extractions and (b) by solubilization of the wall with the pneumococcal autolytic enzyme followed by fractionation of the enzyme products. Two major macromolecular components could be identified. One, the peptidoglycan, is composed of glucosamine, muramic acid (plus muramic acid phosphate), lysine, alanine, and glutamic acid in the molar ratios of 1.0:1.2:1.5:3.8:2.0. The presence of additional amino acids, namely, aspartic acid, serine, glycine, and threonine in the molar ratios (with respect to glucosamine) of 0.9:1.0:1.0:0.5 was also detected. The other major component, teichoic acid, is rich in galactosamine, phosphate, and choline which occur in the molar ratios 1.0:1.65:0.9. These three constituents together make up 23% of the cell wall mass. After treatment of cell walls with cell-free pneumococcal autolytic enzyme, most of the cell wall material can be recovered in the form of two soluble macromolecular fractions which are separable by gel filtration. Fraction I is of high molecular weight and contains teichoic acid polymers plus what appears to be the polysaccharide backbone of the peptidoglycan. Fraction II contains material of lower molecular weight and is rich in the amino acids lysine, glutamic acid, and alanine; these amino acids appear to represent cross-linked dimers and trimers of the peptide portion of the peptidoglycan. The teichoic acid and the polysaccharide components of Fraction I can be separated after sequential degradation with nitrous acid and periodate. It has already been proposed in the literature that the antigenic C-polysaccharide of pneumococcus is a teichoic acid. Our studies further indicate that this teichoic acid contains choline and is a major structural component of pneumococcal cell wall. The major autolytic activity in pneumococcal extracts appears to be an amidase which splits the bond between muramic acid and alanine in the peptidoglycan portion of the cell wall. Experiments are described which indicate that the lysis of pneumococci by deoxycholate occurs through the participation of this enzyme. The choline component of the teichoic acid plays a key role in determining sensitivity to the autolytic enzyme, since walls prepared from pneumococci in which cell wall choline was replaced by ethanolamine were found to be totally resistant to the action of the autolytic enzyme.