Increasing the conformational stability by replacement of heme axial ligand in c‐type cytochrome

FEBS Letters

Kurihara, Masaaki¹  Satoh, Tadashi⁴  Suruga, Kohei⁴  Yamada, Seiji⁴  Itoga, Akito⁴  Fukazawa, Chikafusa²  Suzuki, Noriko⁴  Oku, Tadatake⁴  Arahira, Masaomi²  Natori, Miwa⁴  Nishio, Toshiyuki⁴  Isogai, Yasuhiro³  Kawachi, Ryu⁴ 

[1] National Institute of Health Sciences, Kamiyoga 1-18-1, Setagaya, Tokyo 158-8501, Japan;Genetic Engineering Laboratory, National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan;RIKEN, Hirosawa 2-1, Wako, Saitama 351-0198, Japan;Department of Biological Chemistry, College of Bioresource Sciences, Nihon University, Kameino 1866, Fujisawa, Kanagawa 252-8510, Japan

关键词: Axial ligand;    Conformational stability;    Cytochrome c 6;    Heme iron;    Porphyra yezoensis;    CD;    circular dichroism;    E m;    midpoint redox potential;    non-mutant;    recombinant protein without any mutation;    PCR;    polymerase chain reaction;   

DOI  :  10.1016/S0014-5793(02)03615-3

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

To investigate the role of the heme axial ligand in the conformational stability of c-type cytochrome, we constructed M58C and M58H mutants of the red alga Porphyra yezoensis cytochrome c ₆ in which the sixth heme iron ligand (Met58) was replaced with Cys and His residues, respectively. The Gibbs free energy change for unfolding of the M58H mutant in water (ΔG°_unf=1.48 kcal/mol) was lower than that of the wild-type (2.43 kcal/mol), possibly due to the steric effects of the mutation on the apoprotein structure. On the other hand, the M58C mutant exhibited a ΔG°_unf of 5.45 kcal/mol, a significant increase by 3.02 kcal/mol compared with that of wild-type. This increase was possibly responsible for the sixth heme axial bond of M58C mutant being more stable than that of wild-type according to the heme-bound denaturation curve. Based on these observations, we propose that the sixth heme axial ligand is an important key to determine the conformational stability of c-type cytochromes, and the sixth Cys heme ligand will give stabilizing effects.

【 授权许可】

Unknown