Alterations in the phospholipid head group composition of most strains of Rhodopseudomonas sphaeroides, as well as Rhodopseudomonas capsulata and Paracoccus denitrificans, occurred when cells were grown in medium supplemented with Tris. Growth of R. sphaeroides M29-5 in Tris-supplemented medium resulted in the accumulation of N-acylphosphatidylserine (NAPS) to as much as 40% of the total whole-cell phospholipid, whereas NAPS represented approximately 28 an 33% of the total phospholipid when R. capsulata and P. denitrificans respectively, were grown in medium containing 20 mM Tris. The accumulation of NAPS occurred primarily at the expense of phosphatidylethanolamine in both whole cells and isolated membranes of R. sphaeroides and had no detectable effect on cell growth under either chemoheterotrophic or photoheterotrophic conditions. Yeast extract (0.1%) and Casamino Acids (1.0%) were found to be antagonistic to the Tris-induced (20 mM) alteration in the phospholipid composition of R. sphaeroides. The wild-type strains R. sphaeroides 2.4.1 and RS2 showed no alteration in their phospholipid composition when they were grown in medium supplemented with Tris. In all strains of Rhodospirillaceae tested, as well as in P. denitrificans, NAPS represented between 1.0 and 2.0% of the total phospholipid when cells were grown in the absence of Tris. [32P]orthophosphoric acid entered NAPS rapidly in strains of R. sphaeroides that do (strain M29-5) and do not (strain 2.4.1) accumulate this phospholipid in response to Tris. Our data indicate that the phospholipid head group composition of many Rhodospirillaceae strains, as well as P. denitrificans, is easily manipulated; thus, these bacteria may provide good model systems for studying the effects of these modifications on membrane structure and function in a relatively unperturbed physiological system.