Isolation and Study of the Composition of a Peptidoglycan Complex Excreted by the Biotin‐Requiring Mutant of Brevibacterium divaricatum NRRL‐2311 in the Presence of Penicillin

When cultures of biotin‐requiring mutant of Brevibacterium divaricatum NRRL‐2311 were treated in early logarithmic phase with penicillin (2–3 units/ml) in a glucose‐mineral medium under excess supply of biotin, large amounts of a high‐molecular‐weight material accumulated in the medium. The phenomenon could not be observed with cultures run in parallel from which penicillin was omitted. Chemical analyses of the excreted material isolated from the media of 1‐h and 24‐h penicillin‐treated cultures, showed that the main constituents were peptidoglycan components of noncross‐linked structure bearing both l and d‐alanine residues; evidence was also obtained for the occurrence of extractable lipid material, non‐amino sugars and organic phosphate. Under identical conditions, the excretion of peptidoglycan could be induced by ampicillin and cloxacillin, respectively, but not by bacitracin. Addition of penicillin to biotin‐requiring mutant of M. glutamicus yielded similar results, indicating that the phenomenon was not restricted to the Brev. divaricatum strain. This suggests that excretion of peptidoglycan material by the two biotin‐requiring mutants might be the result of two events, (a) a change in the osmotic barrier of the cell and (b) specific inhibition of cross‐link formation in peptidoglycan induced by penicillin. Several procedures were examined for the isolation and purification of the peptidoglycan complex excreted by penicillin‐treated Brev. mutant. Suitable labelling experiments with l‐[U‐14C]glutamic acid and analyses of lysozyme digests of the purified fractions, suggest that some of the components might contain murein fragments covalently linked to a polysaccharide portion.