Cloning and characterization of a Nicotiana tabacum methylputrescine oxidase transcript

The oxidative deamination of N-methylputrescine is an essential step in both pyridine and tropane alkaloid biosynthesis. Reverse genetic approaches have not resulted in the cloning of a methylputrescine oxidase gene (MPO). However, we have used a homology-based approach to clone a full-length tobacco MPO1 cDNA. The MPO1 gene is part of a small multigene family comprised of approximately six members. MPO1-like transcript levels increased in roots that were either deprived of auxin or treated with methyl jasmonic acid. Similar to other known nicotine biosynthetic genes in domesticated tobacco, MPO1-like mRNA levels were lower in roots with the mutant a and b alleles. The MPO1 protein was expressed in bacteria as a recombinant Thioredoxin-His(6)-MPO1 fusion protein. The recombinant MPOI protein utilized N-methylputrescine more efficiently than other diamines. Therefore, the kinetic properties of the MPOI enzyme may play an important role in determining the pyridine alkaloid profiles observed in tobacco roots. (c) 2006 Elsevier Ltd. All rights reserved.