Capsular polysaccharide (CPS) is a major virulence determinant for human pathogenic bacteria. Although the essential functional roles for CPS in bacterial virulence have been established, knowledge of how CPS production is regulated remains limited. Streptococcus pneumoniae (pneumococcus) CPS expression levels and overall thickness change in response to available oxygen and carbohydrate. These nutrients in addition to transition metal ions can vary significantly between host environmental niches and infection stage. Since pneumococcus must modulate CPS expression among various host niches during disease progression, we examined the impact of nutritional transition metal availability of manganese (Mn) and zinc (Zn) on CPS production. We demonstrate that increased Mn/Zn ratio increase CPS production via Mn-dependent activation of the phosphoglucomutase Pgm, an enzyme that functions at the branch point between glycolysis and the CPS biosynthetic pathway in a transcription-independent manner. Further, we find that the downstream CPS protein CpsB, a Mn-dependent phosphatase, does not promote aberrant dephosporylation of its target capsule-tyrosine kinase CpsD during Mn-stress. Together, these data reveal a direct role for cellular Mn/Zn ratios in the regulation of CPS biosynthesis via direct activation of Pgm. We propose a multilayer mechanism used by the pneumococcus in regulating CPS levels across various host niches. IMPORTANCE Evolving evidence strongly indicates that maintenance of metal homeostasis is essential for establishing colonization and continued growth of bacterial pathogens in the vertebrate host. In this study, we demonstrate the impact of cellular manganese/zinc (Mn/Zn) ratios on bacterial capsular polysaccharide (CPS) production, an important virulence determinant of many human pathogenic bacteria, including Streptococcus pneumoniae. We show that higher Mn/Zn ratios increase CPS production via Mn-dependent activation of the phosphoglucomutase Pgm, an enzyme that functions at the branch point between glycolysis and the CPS biosynthetic pathway. The findings provide a direct role for Mn/Zn homeostasis in the regulation of CPS expression levels and further support the ability for metal cations to act as important cellular signaling mediators in bacteria.

A chemistry course designed as an elective for chemistry, biochemistry, biology, and pharmaceutical science majors is described. The course is designed for second- through fourth-year undergraduate students. The prerequisite is first-semester, second-year undergraduate organic chemistry with the second semester of the organic sequence as a corequisite. The emphasis is on the mechanistic organic chemistry of enzyme transformations. An inductive approach is utilized in which students are initially taught an array of mechanistic probe experiments and learn how to design and implement these experiments to elucidate mechanisms. The intent of this research/hypothesis-driven approach is to enable students to propose experiments to test premises. After proposing probe experiments, the students will receive data so that, upon their interpretation of the data, they can propose potential mechanism(s) and defend their proposals. The primary chemical literature is emphasized throughout the course. Potential syllabi, a detailed overview of the applicable mechanistic probe experiments, multiple problem sets for distribution to students (with a faculty answer guide), a listing of 40 appropriate primary literature research articles utilized in the course, a listing of additional resources, and an extensive student study guide (reviewing applicable organic chemistry and biochemistry review principles) are all provided in the Supporting Information.