Catalyzes the oxidation of glycolate to glyoxylate, with a reduction of O2 to H2O2 (PubMed:1324737, PubMed:7705356). Is a key enzyme in photorespiration in green plants (Probable). To a lesser extent, is also able to use L-lactate and 2-hydroxbyutanoate as substrate in vitro, but shows almost no activity with L-mandelate (PubMed:7705356). glycolate + O2 = glyoxylate + H2O2 a (2S)-2-hydroxycarboxylate + O2 = a 2-oxocarboxylate + H2O2 Binds 1 FMN per subunit. kcat is 20 sec(-1) with glycolate as substrate. kcat is 11 sec(-1) with L-lactate as substrate. kcat is 13.2 sec(-1) with L-2-hydroxbyutanoate as substrate. kcat is 0.09 sec(-1) with L-mandelate as substrate. Photosynthesis; photorespiration; glycine from 2-phosphoglycolate: step 2/3. Homotetramer. Belongs to the FMN-dependent alpha-hydroxy acid dehydrogenase family.