A pyridoxal 5′-phosphate-dependent Mannich cyclase

Pyridoxal 5′-phosphate (PLP)-dependent enzymes catalyse a diverse range of chemical transformations. Despite their extraordinary functional diversity, no PLP-dependent enzyme is known to catalyse Mannich-type reactions, an important carbon–carbon bond-forming reaction in synthetic organic chemistry. Here we report the discovery of a biosynthetic enzyme LolT, a PLP-dependent enzyme catalysing a stereoselective intramolecular Mannich reaction to construct the pyrrolizidine core scaffold in loline alkaloids. Importantly, its versatile catalytic activity is harnessed for stereoselective synthesis of a variety of conformationally constrained α,α-disubstituted α-amino acids, which bear vicinal quaternary–tertiary stereocentres and various aza(bi)cyclic backbones, such as indolizidine, quinolizidine, pyrrolidine and piperidine. Furthermore, crystallographic and mutagenesis analysis and computational studies together provided mechanistic insights and structural basis for understanding LolT’s catalytic activity and stereoselectivity. Overall, this work expands the biocatalytic repertoire of carbon–carbon bond-forming enzymes and increases our knowledge of the catalytic versatility of PLP-dependent enzymes. Pyridoxal 5′-phosphate (PLP)-dependent enzymes that catalyse Mannich reactions were unknown. Now, it is reported that the PLP-dependent enzyme LolT catalyses a 5-endo-trig Mannich cyclization reaction during the pyrrolizidine core scaffold formation in loline biosynthesis, and its crystal structure is solved.