Biologically Active Components from Mycobacterial Cell Walls. I. Isolation and Composition of Cell Wall Skeleton and Component P 3

Abstract

The BCG cell wall was fractionated to isolate and characterize the components responsible for tumor suppression and regression. Treatment of cell walls with proteolytic enzymes, followed by extraction with organic solvents, isolated a soluble fraction designated “free lipids” and an insoluble residue designated “cell wall skeleton” (CWS-I). The latter, a polymeric mycolic acid-arabinogalactan-mucopeptide complex, contained 34% mycolic acid(s), 38.6% arabinogalactan with arabinose and galactose in a molar ratio of 2.8:1, and 20.1% amino acids, similar to those reported for mucopeptides of the mycobacterial cell wall. About 1% of 5 additional amino acids also present presumably were protein remnants not removed by treatment with enzymes. Since CWS-I was as effective as the original cell wall in suppressing tumor growth but did not cause regression of established tumors, we studied conventional wax D and cord factor preparations (subfractions of the free lipids). From both products, a chromatographically pure component, P3 , was isolated by pressure elution through columns of microparticulate silica gel packed by centrifugation. P3 contained trehalose, mycolic acid, and mycolic acids of unknown structure. When P3 was recombined with CWS-I, tumor-regressing activity equal to that of the original cell wall was restored.