To fortify their cytoplasmic membrane and protect it from osmotic rupture, most bacteria surround themselves with a peptidoglycan (PG) exoskeleton. The PG synthases that build this structure are called penicillin-binding proteins (PBPs). Since they are the targets of penicillin and related antibiotics, the structures and in vitro biochemical functions of the PBPs have been extensively studied. However, the in vivo functions of the PBPs and the factors they work with to build the PG meshwork remain poorly understood. PBPs work in the context of multicomponent complexes organized by cytoskeletal elements. A major outstanding question has been whether or not these complexes contain factors required for PBP function. I addressed this using Escherichia coli as a model system by taking advantage of the synthetic lethal phenotype resulting from simultaneous inactivation of the major PG synthases: PBP1a and PBP1b. Using a screen for mutants synthetically lethal with the inactivation of PBP1b, I identified LpoA as a factor required for PBP1a function. A colleague in the lab performed the analogous screen for mutants synthetically lethal with the inactivation of PBP1a and identified LpoB as a factor required for PBP1b function. We showed that the Lpo factors are outer membrane lipoproteins that form specific trans-envelope complexes with their cognate PBPs in the inner membrane and that LpoB can stimulate the activity of PBP1b in vitro. Our results reveal unexpected complexity in the control of PBP activity and indicate that they likely receive regulatory input from the outer membrane in addition to cytoskeletal elements in the cytoplasm. To investigate the role of LpoB in morphogenesis further, I took a genetic approach that has identified PBP1b* variants capable of functioning in vivo in the absence of LpoB. Preliminary characterization of these variants indicates that LpoB has cellular functions in addition to PBP1b activation and that LpoB may be important for coordinating the two different catalytic activities of PBP1b. Future study of these mutants is likely to uncover important insights into PBP function and their control by the Lpo factors. These insights may open new avenues for the development of novel therapeutics that target the PBPs.