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For a long time, amino acid residues have been thought to provide backbone which holds the protein active site in place and directs substrates/products in specific orientations to catalyze chemical reactions. However, ample evidence has appeared in recent literature which signifies the role of redox-active amino acid residues like tyrosine or tryptophan in enzyme catalysis (Chem. Rev. 1998, 98, 705). Such new information has greatly improved our understanding of reaction rates and enzyme catalysis in modern biochemistry.
I will present recent results on two different classes of enzymes: heme containing Rice-α-oxygenase (RαO) from Oryza sativa and Cu-containing small laccase (SLAC) from Streptomyces coelicolor which form tyrosyl radicals during turnover. RαO catalyzes the α-oxidation of long chain fatty acids whereas laccases catalyzes the oxidation of phenols, aromatic amines, etc. I have made use of a variety of spectroscopic methods and enzyme kinetics to study the mechanism of operation of these two enzymes. For RαO, I have demonstrated reversible H• abstraction of the substrate by a Y379• that is formed during enzyme turnover. A very large, weekly temperature dependent kinetic isotope effect (~50) has been observed which is consistent with nuclear tunneling. For SLAC, I have shown that Y108 residue, which resides close to the trinuclear Cu cluster, gets oxidized when there is a shortage of reducing equivalents in the milieu. It is proposed that such reversible oxidation of key protein residues is (one of) the defense mechanisms of cells to prevent oxidative damage of critical machinery by the reactive oxygen species generated during O2 metabolism by the enzymes.
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