A unique AA5 alcohol oxidase fused with a catalytically inactive CE3 domain from the bacterium Burkholderia pseudomallei

Copper radical oxidases (CROs) from Auxiliary Activities family 5 can oxidize various alcohol substrates. Bacterial CROs are less studied than eukaryotic enzymes. Here we have characterized an unusual bacterial CRO that acts only on primary alcohols and is fused to a carbohydrate esterase domain, which was completely inactive due to substitutions of key amino acid residues in the active site. Copper radical oxidases (CROs) are redox enzymes able to oxidize alcohols or aldehydes, while only requiring a single copper atom as cofactor. Studied CROs are found in one of two subfamilies within the Auxiliary Activities family 5 (AA5) in the carbohydrate-active enzymes database. We here characterize an AA5 enzyme outside the subfamily classification from the opportunistic bacterial pathogenBurkholderia pseudomallei, which curiously was fused to a carbohydrate esterase family 3 domain. The enzyme was shown to be a promiscuous primary alcohol oxidase, with an activity profile similar to enzymes from subfamily 2. The esterase domain was inactive on all tested substrates, and structural predictions revealed this being an effect of crippling substitutions in the expected active site residues.
Source: FEBS Letters - Category: Biochemistry Authors: Tags: Research Letter Source Type: research