Partial purification and characteristics of membrane-associated NAD(+)-dependent isocitrate dehydrogenase activity from etiolated pea mitochondria

NAD(+)-dependent isocitrate dehydrogenase from pea mitochondrial matrix has been isolated, partially purified, and characterized [1]. Of the total NAD-ICDH activity found in pea mitochondria, 35-50% is found tightly associated with the membrane. Isolation and partial purification of membrane-associated ICDH showed that this activity has physical properties that differ from those of the matrix form. Membrane-ICDH was not removed from the membrane by washing with weak detergent, conditions which result in solubilization of over 90% of the malate dehydrogenase activity. Both forms of the enzyme bind to Matrex Gel Blue A in the absence of detergent, however 4 x higher salt levels are required to elute the membrane form; membrane-ICDH also binds to Blue A in the presence of detergent while the matrix form does not. Membrane-ICDH is also more anionic requiring higher salt for elution from Mono Q, and in the absence of detergent is unrecoverable from this column. Characterization of ca. 40-fold purified membrane-ICDH revealed that the membrane and matrix form of the enzyme have similar kinetic properties. The possible nature of physical differences between these activities as well as potential physiological roles is discussed. (C) 1997 Elsevier Science Ireland Ltd.