Studies on the ‘incorporation factor’ with Bacillus megaterium

Abstract

The development and details of a simple and reproducible assay for the "incorporation factor" are described. It is based on the stimulation of [14C]glycine incorporation into intact Bacillus megaterium cells by the incorporation factor described by Gale and Fokes (1958). The characteristics of the assay show marked similarities to those of the disrupted Staphylococcus aureus system in the following respects: activity of nucleic acids from B. megaterium and S. aureus and inactivity of commercial nucleic acid preparations; inhibition by 4,5,6-trichlorobenzimidazole; activity of ribonucleosides and inactivity of ribose, free bases and ribonucleotides. A further improvement over the disrupted staphylococcal system lies in the observation that no significant portion of the glycine incorporated by the intact cells of B. megaterium enters the cell-wall fraction. Of a large number of known compounds tested; only glycerol, glucose and fructose were able to replace the incorporation factor completely. On a weight basis the most active compound was glycerol. Permeability difficulties may account for the inactivity of many of the compounds. The activity of nucleic acid prepared from B. megaterium appears to be due to bound glycerol which can be removed by dialysis but not by extraction with ethanol.