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Medientyp:
E-Artikel
Titel:
The Replacement of Trp392 by Alanine Influences the Decarboxylase/Carboligase Activity and Stability of Pyruvate Decarboxylase from Zymomonas mobilis
Beschreibung:
<jats:p>The bulky tryptophan residue 392 located in the deep cleft leading to the active center of pyruvate decarboxylase (PDC) from <jats:italic>Zymomonas mobilis</jats:italic> was changed to alanine which is found in the equivalent position of PDC from yeast. The mutation reduced the decarboxylase activity towards pyruvate by a factor of two (60–70 U/mg), whereas the <jats:italic>K</jats:italic><jats:sub>m</jats:sub> (1.1 mM in Mes/KOH buffer) remains unchanged compared with the wild‐type enzyme. The apparent <jats:italic>K</jats:italic><jats:sub>m</jats:sub> for thiamine diphosphate (thiamin–<jats:italic>P</jats:italic><jats:sub>2</jats:sub>) in the presence of 5 mM MgSO<jats:sub>4</jats:sub> was increased by a factor of 10 (84 μM in Mes/KOH buffer) and the tetrameric mutant protein was less stable, as indicated by urea denaturation experiments. The mutation enhanced the carboligase activity of the enzyme towards benzaldehyde by a factor of four. The resulting α‐hydroxyketone was identified as (<jats:italic>R</jats:italic>)‐phenylacetylcarbinol.</jats:p>