C-13 NUCLEAR MAGNETIC-RESONANCE STUDIES OF THE INTERACTION OF LECITHIN WITH PURIFIED D-BETA-HYDROXYBUTYRATE APODEHYDROGENASE, A LIPID-REQUIRING ENZYME

The interaction of d-β-hydroxybutyrate apodehydrogenase (BDH), a lecithin-requiring enzyme purified from bovine heart mitochondria, with lecithin (PC) has been studied by 13C NMR by using lecithin labeled either in the polar moiety, soybean [N-13CH3]PC, or in the hydrophobic moiety, [11-13C]dioleoyl-PC. The lecithin was admixed together with phosphatidylethanolamine and diphosphatidylglycerol, the other two major mitochondrial phospholipids and added as vesicles in aqueous buffer. With increasing BDH/PC ratio, he spin-lattice relaxation time (T1) of the choline N-13CH3 signal (resonance at 54 ppm) decreased, whereas T1 increased for the hydrophobic label, [11-13C]dioleoyl-PC (resonance at 27 ppm). For relaxation by a dipole-dipole interaction mechanism, 1/T1 is proportional to tc, the rotational correlation time of the 13C nucleus, so that changes in 1/T1 reflect changes in the rotational motion of the 13C-labeled moiety. At the highest protein to lipid ratio studied, 4,8 mg of BDH added per mg of PC, 1/T1 of the N-13CH3 signal was enhanced 60% compared with that of phospholipid vesicles alone, whereas for the 11-13C resonance 1/T1 was diminished by 33%. These studies show that, upon interaction of BDH with phospholipid vesicles containing lecithin, the rotational motion in the polar group of lecithin is constrained, whereas the motion in the hydrophobic region is increased. The increased motion of the hydrophobic moiety could result from disorder in the bilayer. Futher, the line shape of the 11-13C lecithin resonance was broadened upon interaction with BDH. Such line broadening could result from chemical shift anisotropy or constrained lateral motion. The interaction of BDH with lecithin in phospholipid vesicles, as measured by 13C NMR, is unique as compared with the two other systems previously studied. © 1979, American Chemical Society. All rights reserved.