Sequence-based screening and characterization of cytosolic mandelate oxidase using oxygen as electron acceptor

Sequence-based screening was carried out to find a type of cytosolic mandelate oxidase that converted l-mandelate to phenylglyoxylate using oxygen as the final electron acceptor. The sequence features of the cytosolic mandelate oxidase were summarized, and were used in the screening process. Mandelate oxidases from Streptomyces coelicolor (Hmo_SC) and Amycolatopsis orientalis (Hmo_AO) were screened and then they were heterologously expressed and characterized. At pH 7.3 40 °C, the Hmo_AO showed k_cat and K_m values of 140 min⁻¹ and 10.2 mM, the Hmo_SC showed k_cat and K_m values of 105.1 min⁻¹ and 2.06 mM. The Hmo_SC was thermal stable and retained its 90% activity at 60 °C for up to 5 h, while Hmo_AO lost most of its activity at this temperature. The Hmo_SC could effectively catalyze the conversion of l-mandelate to phenylglyoxylate at higher temperature using oxygen as the final electron acceptor.