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The rates of solvent exchange for the C5 and C3 sites of the TPQ cofactor in wild-type amine oxidases and its mutants from Hansenula polymorpha are discussed. Data reveal that a rapid rate of C5=O exchange and a slow to zero rate of C3-H exchange suggest multiple orientations for the TPQ cofactor.

Topaquinone formation in a yeast copper amine oxidase was found to be dependent on an intact copper binding site, in studies using a heterologous expression systems and site-directed mutagenesis. Topaquinone was fully formed in the E406N mutant of methylamine oxidase, indicating that the negatively charged residue at the +1 position is not essential to modification of the tyrosyl side chain.

Resonance Raman (RR) spectroscopy is a useful method of providing structural information about the 2,4,5-trihydroxyphenylalaniequinone (TPQ) cofactor, for identifying the source of oxygen atoms during synthesis. The C=O stretch f the C5 carbonyl at 1683 cm-1 and the C=O stretch of the C2 carbonyl at 1575 cm-1 was identified by specific labeling of the C2, C4 and C5 oxygens of TPQ from Arthrobacter globiformis. C-O at C5 showed greater double-bond character than at C2 or C4. Bovine serum amine oxidase showed a similar nu(C=O) mode at 1678 cm-1.

The state of the carbonyl group methylamine hydrogenase and its endogenous tryptophan tryptophylquinone (TTQ) factor is investigated using resonance Raman spectroscopy. Results indicate a strong electrostatic interaction between monovalent cations and a carbonyl oxygen in TTQ. The direct electrostatic interaction between the monovalent cations and the TTQ factor explain the ability of the former to promote and stabilize the formation of the semiquinone intermediate.

A highly purified enzyme was used to obtain the resonance Raman (RR) spectra of the 2,4,5-trihydroxyphenylalanine quinone cofactor in Escherichia coli amine oxidase from strain W3350. The substrate amine group was observed to be bound to the cofactor in the semiquinone state. In addition, Cu(I) cyanide complex formation was found to perturb the topa(sub NHSQ)'s absorption and RR spectra.

X-ray diffraction studies of copper(II) complexes containing coenzyme PQQ and terpyridine revealed the influence of the pyrrole moiety on the PQQ molecule. Pyrrole moiety facilitated the coordination between the PQQ molecule and Cu(II), while quinone moiety disallowed the binding of PQQ molecule to Cu(II). The O,O-coordination of quinone moiety was observed in the PQQ complex.

The reaction of (Cu bpy H2O 2)(BF4)2 with DL-topa in 1:1 ratio under Ar atmosphere produced (Cu DL-topa bpy H2O)BF4.3H2O crystals. The X-ray analysis revealed the preparation of an (N,O) complex between the amino and carboxyl groups which promoted the formation of a square pyramidal configuration for the Cu(II) ion. Furthermore, an intramolecular stacking occurred between the aromatic ring of the topa and the bpy ring. The crystal structure featured an extensive system of hydrogen-bonding interactions. In addition, the product induced the oxidation of benzylamine at room temperature under aerobic conditions.

Single site-directed mutants F310A and A320Q and double mutant F310A/A320Q are constructed and all mutant enzymes as well as wild-type (WT) cytochrome P450 from the thermoacidophilic crenarchaeon Sulfolobus tokodaii strain 7 (P450st) are expressed at high levels. The results have shown that the reduction potential and electron density on the heme iron are modulated by the Phe310 and Ala320 residues in P450st.