Theoretical study of the conformation of the lipoamide arm in a mutant H protein

The lipoamide arm of the H protein plays a pivotal role in the catalytic cy cle of the glycine decarboxylase complex (GDC) by being successively methyl amine loaded (Hmet), reduced (Hred), and oxidized (Hox). In a previous stud y, we calculated free-energy surfaces as a function of the lipoamide arm po sition of the three forms of the wild-type protein and found close agreemen t with the available experimental data. Our simulations, together with crys tallographic and NMR data, showed that the methylamine-loaded arm is locked in a cavity by interaction with Ser12, Glu14, and Asp67. In this work, we investigate the behavior of the methylamine-loaded form of a mutant H prote in (HEA) where Glu14 has been replaced by Ala. We find that the arm can sti ll be held in the cavity but that the energy barrier to release of the arm is halved from similar to 40 kcal mol(-1) for Hmet to similar to 12 kcal mo l(-1) for HEA. To compensate for the loss of Glu14, the methylamine group s hifts toward Ser66 in the mutant form. These results provide a structural b asis for the equilibrium between the loaded and the unloaded forms of the a rm observed by Gueguen et al. (Gueguen et al., J Biol Chem 1999;274:2634426 352) in HEA. (C) 2001 Wiley-Liss, Inc.