Toxic at high concentrations, H2S is a signaling molecule produced by cells and modulates important physiological processes including blood pressure regulation, inflammation and neuormodulation. Our laboratory is investigating the reaction mechanisms and regulation of enzymes involved in H2S biogenesis and its clearance via oxidation. In addition to the canonical mitochondrial sulfide oxidation pathway, we have recently discovered a new pathway for clearing H2S, which involves hemeproteins. We use a combination of spectroscopic (EPR, fluorescence), kinetic (stopped-flow spectroscopy) and cellular approaches to understand the mechanisms of catalysis and regulation of key enzymes involved in H2S homeostasis. The enzymes involved in sulfur metabolism are richly dependent on multiple B vitamins for their catalytic functions including vitamin B6, folic acid and B12. My laboratory is also studying the intricate network of chaperones that shepherd and tailor vitamin B12 from its point of entry into cells to its target enzymes and whose dysfunction lead to disease. We have been elucidating novel enzymatic functions of the individual proteins and the thermodynamics and kinetics of protein-protein interactions in the pathway that guide B12 delivery without dilution into the cellular milieu. Using a combination of structural, spectroscopic and kinetic approaches we are studying allosteric regulation in the trafficking pathway for cofactor delivery with high fidelity.