Studies of lysine cyclodeaminase from Streptomyces pristinaespiralis: Insights into the complex transition NAD+ state.

Lysine cyclodeaminase (LCD) catalyzes the piperidine ring formation in macrolide-pipecolate natural products metabolic pathways from a lysine substrate through a combination of cyclization and deamination. This enzyme belongs to a unique enzyme class, which uses NAD+ as the catalytic prosthetic group instead of as the co-substrate. To understand the molecular details of NAD+ functions in lysine cyclodeaminase, we have determined four ternary crystal structure complexes of LCD-NAD+ with pipecolic acid (LCD-PA), lysine (LCD-LYS), and an intermediate (LCD-INT) as ligands at 2.26-, 2.00-, 2.17- and 1.80 Å resolutions, respectively. By combining computational studies, a NAD+-mediated "gate keeper" function involving NAD+/NADH and Arg49 that control the binding and entry of the ligand lysine was revealed, confirming the critical roles of NAD+ in the substrate access process. Further, in the gate opening form, a substrate delivery tunnel between ε-carboxyl moiety of Glu264 and the α-carboxyl moiety of Asp236 was observed through a comparison of four structure complexes. The LCD structure details including NAD+-mediated "gate keeper" and substrate tunnel may assist in the exploration the NAD+ function in this unique enzyme class, and in regulation of macrolide-pipecolate natural product synthesis.