Gram-negative bacteria are enclosed by two membranes with different compositions. The outer membrane normally has LPS, a lipid with sugars attached to it. We recently discovered that a model bacterium can survive without LPS if it produces CPG2, a type of ceramide lipid. This shows the importance of ceramides in bacterial biology and provides an opportunity to study alternative modes of outer membrane construction in bacteria. The proposed research is anticipated to discover how CPG2 is made and explain how the enzymes that make it work. This knowledge will be essential to discover the numerous roles of bacterial lipids in the various environments in which bacteria live, including the human body. Because many types of bacteria produce ceramides, this advance will contribute to answering important basic and translational research questions related to the production and function of lipids in all bacteria, including those causing disease. More broadly, this project will train students at the high school, undergraduate, and graduate levels. Specifically, these students will be trained in a multidisciplinary setting to develop skills in genetics, biochemistry, and structural biology. These skills are essential to develop the new generation of researchers, and a strong biotech workforce. Gram-negative bacteria are characterized by an envelope consisting of two membranes. The inner membrane is largely composed of phospholipids and functions similarly to the eukaryotic plasma mem