The high-level heterologous expression in Pichia pastoris, purification and characterization of recombinant membrane-bound rat liver monoamine oxidase A (MAO A) are described. A I-L culture of cells produces similar to 700 U of rat MAO A activity. The rat MAO A activity is found in outer mitochondrial membrane of the cell. Using a modification of the human MAO A purification procedure, similar to 200 mg of recombinant rat MAO A is purified in a 43% yield and exhibits a molecular weight of similar to 60,000 kDa on SDS-PAGE. The purified enzyme contains a covalently bound FAD and forms a N(5) flavocyanine adduct on inhibition by clorgyline. Edman sequencing shows that the amino terminus of rat MAO A is blocked at an N-terminal threonyl residue. The purified rat enzyme exhibits a higher thermal stability than does purified human MAO A. Compared with human MAO A, rat MAO A oxidizes serotonin or kynuramine with twofold higher k(cat)/K-m values, oxidizes phenethylamine with a 6.7-fold higher catalytic efficiency and benzylamine with a similar to 40-fold higher catalytic efficiency. Although similar to 90% identical in sequence to human MAO A, rat MAO A is a more efficient catalyst for amine neurotransmitter oxidation. (C) 2009 Elsevier Inc. All rights reserved.