Structural Basis for β-Carboline Alkaloid Production by the Microbial Homodimeric Enzyme McbB

The beta-carboline (beta C) alkaloids occur throughout nature and exhibit diverse biological activities. In contrast to beta C alkaloid synthesis in plants, the biosynthesis in microorganisms remains poorly understood. The recently reported McbB from Marinactinospora thermotolerans is a novel enzyme proposed to catalyze the Pictet-Spengler (PS) reaction of L-tryptophan and oxaloacetaldehyde to produce the beta C scaffold of marinacarbolines. In this study, we solved the crystal structure of McbB complexed with L-tryptophan at 2.48 angstrom resolution, which revealed the novel protein folding of McbB and the totally different structure from those of other PS condensation catalyzing enzymes, such as strictosidine synthase and norcoclaurine synthase from plants. Structural analysis and site-directed mutagenesis confirmed that the previously proposed catalytic Glu97 at the active-site center functions as an acid and base catalyst. Remarkably, the structure-based mutants R72A and H87A, with expanded active-site cavities, newly accepted bulky phenylglyoxal as the aldehyde substrate, to produce 1-benzoyl-3-carboxy-beta-carboline.