Biochimica et Biophysica Acta - General Subjects (2020)
Update date:2022-08-15
Topics:
Godoy, Andre Schutzer
Higasi, Paula Miwa Rabelo
Kadowaki, Marco Antonio Seiki
Polikarpov, Igor
Prade, Rolf Alexander
de Araújo, Evandro Ares
de Godoy, Mariana Ortiz
Background Fungal aryl-alcohol oxidases (AAOx) are extracellular flavoenzymes that belong to glucose-methanol-choline oxidoreductase family and are responsible for the selective conversion of primary aromatic alcohols into aldehydes and aromatic aldehydes to their corresponding acids, with concomitant production of hydrogen peroxide (H2O2) as by-product. The H2O2 can be provided to lignin degradation pathway, a biotechnological property explored in biofuel production. In the thermophilic fungus Thermothelomyces thermophilus (formerly Myceliophthora thermophila), just one AAOx was identified in the exo-proteome. Methods The glycosylated and non-refolded crystal structure of an AAOx from T. thermophilus at 2.6 ? resolution was elucidated by X-ray crystallography combined with small-angle X-ray scattering (SAXS) studies. Moreover, biochemical analyses were carried out to shed light on enzyme substrate specificity and thermostability. Results This flavoenzyme harbors a flavin adenine dinucleotide as a cofactor and is able to oxidize aromatic substrates and 5-HMF. Our results also show that the enzyme has similar oxidation rates for bulky or simple aromatic substrates such as cinnamyl and veratryl alcohols. Moreover, the crystal structure of MtAAOx reveals an open active site, which might explain observed specificity of the enzyme. Conclusions MtAAOx shows previously undescribed structural differences such as a fully accessible catalytic tunnel, heavy glycosylation and Ca2+ binding site providing evidences for thermostability and activity of the enzymes from AA3_2 subfamily. General significance Structural and biochemical analyses of MtAAOx could be important for comprehension of aryl-alcohol oxidases structure-function relationships and provide additional molecular tools to be used in future biotechnological applications.
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