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Structural and Kinetic Properties of the Aldehyde Dehydrogenase NahF, a Broad Substrate Specificity Enzyme for Aldehyde Oxidation
The
salicylaldehyde dehydrogenase (NahF) catalyzes the oxidation
of salicylaldehyde to salicylate using NAD<sup>+</sup> as a cofactor,
the last reaction of the upper degradation pathway of naphthalene
in <i>Pseudomonas putida</i> G7. The naphthalene is an abundant
and toxic compound in oil and has been used as a model for bioremediation
studies. The steady-state kinetic parameters for oxidation of aliphatic
or aromatic aldehydes catalyzed by 6xHis-NahF are presented. The 6xHis-NahF
catalyzes the oxidation of aromatic aldehydes with large <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> values close to 10<sup>6</sup> M<sup>ā1</sup> s<sup>ā1</sup>. The active site
of NahF is highly hydrophobic, and the enzyme shows higher specificity
for less polar substrates than for polar substrates, e.g., acetaldehyde.
The enzyme shows Ī±/Ī² folding with three well-defined domains:
the oligomerization domain, which is responsible for the interlacement
between the two monomers; the Rossmann-like fold domain, essential
for nucleotide binding; and the catalytic domain. A salicylaldehyde
molecule was observed in a deep pocket in the crystal structure of
NahF where the catalytic C284 and E250 are present. Moreover, the
residues G150, R157, W96, F99, F274, F279, and Y446 were thought to
be important for catalysis and specificity for aromatic aldehydes.
Understanding the molecular features responsible for NahF activity
allows for comparisons with other aldehyde dehydrogenases and, together
with structural information, provides the information needed for future
mutational studies aimed to enhance its stability and specificity
and further its use in biotechnological processes