The observation that these 2NTDO point mutations could be associated with substrate and O 2 transport tunnels furthermore supports the notion that an enzymatic reaction can become more efficient while maintaining the kinetic mechanism of enzyme catalysis. The same structural elements have, in fact, been identified as hotspots for engineering Rieske oxygenases not only towards altering C-H hydroxylation selectivity but also for transformation of alternative substrates. The observation that evolving biodegradation potential and rational engineering can be related to amino acid substitution at the same structural entities offers a promising avenue to assign change in Rieske oxygenase structure to its function. This kind of insights will be critical for making more causal connections between structural features of this class of enzymes and the biodegradability of organic soil and water contaminants. We note, however, that it remains to be elucidated how modifications in substrate and O 2 transport tunnels would modulate substrate binding affinities in the active site and the oxygenation efficiencies of target substrates only.