Engineered chimeras reveal the structural basis of hexacoordination in globins: A case study of neuroglobin and myoglobin

I. BORON; L. CAPECE; F. PENNACHIETTI; D. WETZLER; S. BRUNO; S. ABBRUZZETTI; L. CHISARI; F.J. LUQUE; C. VIAPPIANI; M.A. MARTI; D.A. ESTRIN; A. NADRA

BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS

ELSEVIER SCIENCE BV

Lugar: Amsterdam; Año: 2015 vol. 1850 p. 169 - 177

0304-4165

Background: Myoglobin (Mb) and neuroglobin (Ngb) are representative members of pentacoordinated and bis-histidyl, hexacoordinated globins. In spite of their low sequence identity, they show surprisingly similar three-dimensional folds. The ability of Ngb to form a hexacoordinated bis-histidyl complex with the distal HisE7 hasa strong impact on ligand affinity. The factors governing such different behaviors have not been completelyunderstood yet, even though they are extremely relevant to establish structure?function relationships withinthe globin superfamily.Methods: In this work we generated chimeric proteins by swapping a previously identified regulatory segment ?the CD region ? and evaluated comparatively the structural and functional properties of the resulting proteins bymolecular dynamics simulations, and spectroscopic and kinetic investigations.Results: Our results show that chimeric proteins display heme coordination properties displaced towards thoseexpected for the corresponding CD region. In particular, in the absence of exogenous ligands, chimeric Mbis found as a partially hexacoordinated bis-histidyl species, whereas chimeric Ngb shows a lower equilibriumconstant for forming a hexacoordinated bis-histidyl species.Conclusions: While these results confirm the regulatory role of the CD region for bis-histidyl hexacoordination,they also suggest that additional sources contribute to fine tune the equilibrium.General significance: Globins constitute a ubiquitous group of heme proteins widely found in all kingdoms of life.These findings raise challenging questions regarding the structure?function relationships in these proteins, asbis-histidyl hexacoordination emerges as a novel regulatory mechanism of the physiological function of globins.