1MNI

ALTERATION OF AXIAL COORDINATION BY PROTEIN ENGINEERING IN MYOGLOBIN. BIS-IMIDAZOLE LIGATION IN THE HIS64-->VAL(SLASH)VAL68-->HIS DOUBLE MUTANT

1MNI の概要

• エントリーDOI: 10.2210/pdb1mni/pdb
• 分子名称: MYOGLOBIN, PROTOPORPHYRIN IX CONTAINING FE (3 entities in total)
• 機能のキーワード: oxygen storage
• 由来する生物種: Sus scrofa (pig)
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 35199.86

構造登録者

Krzywda, S.,Wilkinson, A.J. (登録日: 1995-01-11, 公開日: 1995-04-20, 最終更新日: 2024-02-14)

主引用文献

Dou, Y.,Admiraal, S.J.,Ikeda-Saito, M.,Krzywda, S.,Wilkinson, A.J.,Li, T.,Olson, J.S.,Prince, R.C.,Pickering, I.J.,George, G.N.
Alteration of axial coordination by protein engineering in myoglobin. Bisimidazole ligation in the His64-->Val/Val68-->His double mutant.
J.Biol.Chem., 270:15993-16001, 1995

PubMed Abstract: Pig and human myoglobin have been engineered to reverse the positions of the distal histidine and valine (i.e. His64(E7)-->Val and Val68(E11)-->His). Spectroscopic and ligand binding properties have been measured for human and pig H64V/V68H myoglobin, and the structure of the pig H64V/V68H double mutant has been determined to 2.07-A resolution by x-ray crystallography. The crystal structure shows that the N epsilon of His68 is located 2.3 A away from the heme iron, resulting in the formation of a hexacoordinate species. The imidazole plane of His68 is tilted relative to the heme normal; moreover it is not parallel to that of His93, in agreement with our previous proposal (Qin, J., La Mar, G. N., Dou, Y., Admiraal, S. J., and Ikeda-Saito, M. (1994) J. Biol. Chem. 269, 1083-1090). At cryogenic temperatures, the heme iron is in a low spin state, which exhibits a highly anisotropic EPR spectrum (g1 = 3.34, g2 = 2.0, and g3 < 1), quite different from that of the imidazole complex of metmyoglobin. The mean iron-nitrogen distance is 2.01 A for the low spin ferric state as determined by x-ray spectroscopy. The ferrous form of H64V/V68H myoglobin shows an optical spectrum that is similar to that of b-type cytochromes and consistent with the hexacoordinate bisimidazole hemin structure determined by the x-ray crystallography. The double mutation lowers the ferric/ferrous couple midpoint potential from +54 mV of the wild-type protein to -128 mV. Ferrous H64V/V68H myoglobin binds CO and NO to form stable complexes, but its reaction with O2 results in a rapid autooxidation to the ferric species. All of these results demonstrate that the three-dimensional positions of His64 and Val68 in the wild-type myoglobin are as important as the chemical nature of the side chains in facilitating reversible O2 binding and inhibiting autooxidation.

PubMed: 7608158
DOI: 10.1074/jbc.270.27.15993

実験手法

X-RAY DIFFRACTION (2.07 Å)