Updated project metadata. BipA is a conserved translational GTPase that resembles elongation factor EF-G and 30S assembly factor LepA. Recent evidence suggests that BipA functions in 50S subunit assembly, but the precise role of the factor remains unclear. Here, we use stable isotope labeling of amino acids in culture and mass spectrometry (SILAC / MS) to examine the function of BipA in ribosome biogenesis. During growth at suboptimal temperature, loss of BipA leads to accumulation of immature large subunit particles (~40S) that lack several proteins. These include L2, L7/12, L10, L14, L16, L17, L19, L27, L28 and L32. Parallel analysis of the control (wild-type) strain shows accumulation of virtually identical intermediate particles, although at much lower levels. Further analysis showed that the main path of 50S assembly differs depending on media in which the cells are grown, demonstrating the robust and flexible nature of the assembly process.