NMR and CD conformational studies of the C-terminal 16-peptides of Pseudomonas aeruginosa c(551) and Hydrogenobacter thermophilus c(552) cytochromes

The 16-amino acid sequences of the C-terminal helices of the homologous bacterial cytochromes C-551 from Pseudomonas aeruginosa and C-552 from Hydrogenobacter thermophilus were synthesized and their solution structure studied. Circular dichroism and NMR experiments in aqueous solution have shown the presence of alpha -helices and 3(10)-helices. The populations of helical structures in phosphate buffer, pH 3.5, 293 K, were 21% for C-551 and 20% for C-552, but increased to 56.7 and 48%, respectively, in 50% aqueous 2,2,2-trifluoroethanol. An isodichroic point was observed at 203 nm in CD spectra for the helix/coil transition in mixtures of water/2, 2,2-trifluoroethanol. NMR spectra in phosphate buffer show the presence of both alpha- and 3(10)-helical structures. In water/2,2,2-trifluoroethanol (50:50) alpha -helices are predominant. CD temperature-dependency studies indicate that both peptides exhibit the same cooperativity for the transition in waterl 2,2,2-trifluoroethanol (50:50). The experimental data show that the amino acid substitutions do not favor heat resistance of the secondary structure of the C-552 C-terminal helix at the local level. Instead, they optimize nonlocal contacts of the polypeptide chain, which stabilize the tertiary structure in the native protein.