Posttranslational biosynthesis of the protein-derived cofactor tryptophan tryptophylquinone

Abstract

Methylamine dehydrogenase (MADH) catalyzes the oxidative deamination of methylamine to formaldehyde and ammonia. Tryptophan tryptophylquinone (TTQ) is the protein-derived cofactor of MADH required for this catalytic activity. TTQ is biosynthesized through the posttranslational modification of two tryptophan residues within MADH, during which the indole rings of two tryptophan side chains are cross-linked and two oxygen atoms are inserted into one of the indole rings. MauG is a c-type diheme enzyme that catalyzes the final three reactions in TTQ formation. In total, this is a six-electron oxidation process requiring three cycles of MauG-dependent two-electron oxidation events using either H2O2 or O2. The MauG redox form responsible for the catalytic activity is an unprecedented bis-Fe(IV) species. The amino acids of MADH that are modified are ≈ 40 Å from the site where MauG binds oxygen, and the reaction proceeds by a hole hopping electron transfer mechanism. This review addresses these highly unusual aspects of the long-range catalytic reaction mediated by MauG.

Figure 1

Protein-derived quinone cofactors. TPQ, 2,4,5-trihydroxyphenylalanine quinone; LTQ, lysine tyrosylquinone; CTQ, cysteine tryptophylquinone; TTQ, tryptophan tryptophylquinone.

Figure 2

Crystal structure of methylamine dehydrogenase (MADH). The overall fold of MADH (PDB code: 2bbk) (6) is represented in cartoon (MADH α-subunit, blue; MADH β-subunit, green) with TTQ drawn in stick colored by atom. Figure produced using PyMOL (www.pymol.org).

Figure 3

The mau gene cluster of P. denitrificans. Of these genes only the gene products of mauB, mauA, mauC and mauG have been isolated and characterized. For the other genes assignment of structure or function is based on sequence similarity to other genes of known function. The information in this figure is based on data presented in references (23, 24, 26).

Figure 4

Reactions catalyzed by MauG that have been monitored by steady-state kinetics. (A) MauG-dependent TTQ biosynthesis from preMADH. (B) MauG-dependent oxidation of reduced quinol MADH to TTQ.

Figure 5

A model for MauG-dependent TTQ biosynthesis from preMADH. As the order of the cross-linking and hydroxylation steps are not known, the two alternative possible routes to form the quinol TTQ are presented. The post-translational modifications are shown in red.

Figure 6

UV-visible absorption spectra of different redox states of MauG. (A) diferric (black) and diferrous (red). (B) Diferric (black) and bis-Fe(IV) (red).

Figure 7

Crystal structures of MauG in complex with precursor methylamine dehydrogenase (preMADH). (A) Overall fold of MauG colored by domain (PDB code: 3l4m) (39). (B) Key components of electron transfer between preMADH and MauG. (C) PreTTQ site before (left panel) and after (right panel) addition of H₂O₂ to MauG-preMADH crystals (PDB codes: 3l4m (left) and 3l4o (right)). The proteins are represented in cartoon (MADH α-subunit, blue; MADH β-subunit, green; panel (B), MauG, pink). Hemes, Trps, TTQ and preTTQ drawn in stick colored by atom (panel (B), carbon, color of associated protein chain cartoon; panel (C), preTTQ, light green, TTQ, dark green). Irons and calcium are drawn as orange and green spheres, respectively. 2Fo-Fc electron density (blue) contoured at 1.0σ. Figure produced using PyMOL (www.pymol.org).

Figure 8

Crystal structure of nitric oxide in complex with diferrous MauG-preMADH (PDB code: 2pxw) (43). NO binds to the heme that lies furthest from the MauG-preMADH interface. Colors and representation are as in Figure 7. Hydrogen bonds are indicated by dashed lines. Figure produced using PyMOL (www.pymol.org).

Figure 9

The preTTQ site of the preMADH β subunit showing Asp residues that are critical for the hydroxylation of βTrp57 to form preTTQ (PDB code: 3l4m) (39). Colors and representation are as in Figure 7. Figure produced using PyMOL (www.pymol.org).

Figure 10

Electron transfer reactions that have been characterized by single-turnover kinetics. (A) The initial two-electron oxidation step in TTQ biosynthesis from preMADH to bis-Fe(IV) MauG. (B) The final two-electron oxidation step in TTQ biosynthesis from quinol MADH to bis-Fe(IV) MauG. (C) The non-biosynthetic electron transfer reaction from diferrous MauG to quinone MADH.