engleski

Brassica rapa auxin-amidohydrolase: the Mn2+ binding affinities and the active site structure

Brassica rapa auxin-amidohydrolase (BrILL2) belongs to the M20D metallopeptidase subfamily, related to the amidohydrolase superfamily (M20) of enzymes which hydrolyze a number of different substrates, including amino acids, sugars, nucleic acids, and organophosphate esters. BrILL2 participates in homeostasis of the plant hormone auxin in a way to hydrolases amino acid conjugates (AACs) of auxins (IAA, IBA, IPA). A large concentration of free auxins, of which the most common is indole-3- acetic acid (IAA), is toxic for plants, so only about 5% of the total concentration of auxin molecules in plants is in the free (active) form, while the rest is stored in inactive forms, mostly as amino acid and sugar conjugates. Auxin amidohydrolases specifically hydrolyze the amide bond of amino acid conjugated auxins (inactive, storage forms), releasing free active compounds. In order to be active BrILL2 needs manganese. The aim of our research was to determine number of Mn2+, in the enzyme active site, and the influence of Cys to Ser mutations on the protein structure and activity. In order to fulfil this aim we conducted an interdisciplinary study combining different experimental and computational approaches: biochemical, spectroscopic, calorimetric and computational.

Brassica rapa auxin-amidohydrolase ; M20 familiy of enzymes ; manganese ions ; auxin ; ITC ; DSC ; computational approach

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