Phenylalanine Ammonia-Lyase-Catalyzed Deamination of an Acyclic Amino Acid: Enzyme Mechanistic Studies Aided by a Novel Microreactor Filled with Magnetic Nanoparticles

Weiser, Diána and Becze, László Csaba and Bánóczi, Gergely and Ender, Ferenc and Kiss, Róbert and Kókai, Eszter and Szilágyi, András Ferenc and Vértessy, Beáta (Grolmuszné) and Farkas, Ödön and Paizs, Csaba and Poppe, László (2015) Phenylalanine Ammonia-Lyase-Catalyzed Deamination of an Acyclic Amino Acid: Enzyme Mechanistic Studies Aided by a Novel Microreactor Filled with Magnetic Nanoparticles. CHEMBIOCHEM, 16. pp. 2283-2288. ISSN 1439-4227

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Abstract

Phenylalanine ammonia-lyase (PAL), found in many organisms, catalyzes the deamination of l-phenylalanine (Phe) to (E)-cinnamate by the aid of its MIO prosthetic group. By using PAL immobilized on magnetic nanoparticles and fixed in a microfluidic reactor with an in-line UV detector, we demonstrated that PAL can catalyze ammonia elimination from the acyclic propargylglycine (PG) to yield (E)-pent-2-ene-4-ynoate. This highlights new opportunities to extend MIO enzymes towards acyclic substrates. As PG is acyclic, its deamination cannot involve a Friedel–Crafts-type attack at an aromatic ring. The reversibility of the PAL reaction, demonstrated by the ammonia addition to (E)-pent-2-ene-4-ynoate yielding enantiopure l-PG, contradicts the proposed highly exothermic single-step mechanism. Computations with the QM/MM models of the N-MIO intermediates from l-PG and l-Phe in PAL show similar arrangements within the active site, thus supporting a mechanism via the N-MIO intermediate.

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