Mitochondria in the White-Rot Fungus Phanerochaete chrysosporium: Purification and Evidence for a Mitochondrial Isoform of Aspartate Aminotransferase

Abstract

Pireaux, J-C., Hayani-Obeidou, W., Chalot, M., Botton, B., and Dizengremel, P. 1995. Mitochondria in the white-rot fungus Phanerochaete chrysosporium: Purification and evidence for a mitochondrial isoform of aspartate aminotransferase. Experimental Mycology 19: 91-100. A very high specific aspartate aminotransferase activity was found in young Phanerochaete chrysosporium cultures. In order to obtain more information about the localization of aspartate aminotransferase, mitochondria were isolated and purified. The purification gradient showed two mitochondrial fractions. Mitochondria localized near the top of the gradient were severely damaged, whereas mitochondria aggregated at the bottom part were largely intact (91 and 94% intactness for outer and inner membrane, respectively). The highest oxidation rates were obtained with succinate and NADH as substrates. Malate oxidation was improved by exogenous NAD⁺ while NADPH oxidation was partially Ca²⁺ dependent. All these oxidations were correctly coupled to the phosphorylation. Experiments using standard respiratory chain inhibitors indicate that, in P. chrysosporium mitochondria, the alternative pathway was present, but not engaged. On nondenaturing uniform polyacrylamide gels, four aspartate aminotransferase isoforms were detected, one being localized in the mitochondrial matrix. On nondenaturing polyacrylamide gradient gels, the nonmitochondrial isoforms had similar electrophoretic mobilities with a molecular mass estimated at 110 kDa, while the mitochondrial isoform had a molecular mass of about 160 kDa.