title: Engineered chimeras reveal the structural basis of hexacoordination in globins: a case study of neuroglobin and myoglobin.
abstract: 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 has a strong impact on ligand affinity. The factors governing such different behaviors have not been completely understood yet, even though they are extremely relevant to establish structure-function relationships within the 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 by molecular dynamics simulations, and spectroscopic and kinetic investigations.
  
  
  RESULTS
  Our results show that chimeric proteins display heme coordination properties displaced towards those expected for the corresponding CD region. In particular, in the absence of exogenous ligands, chimeric Mb is found as a partially hexacoordinated bis-histidyl species, whereas chimeric Ngb shows a lower equilibrium constant 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, as bis-histidyl hexacoordination emerges as a novel regulatory mechanism of the physiological function of globins.
authors.affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina; IQUIBICEN-CONICET, Argentina.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Boron
authors.affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina; Departamento de Química Inorgánica, Analítica y Química Física, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina.
authors.forename: Luciana
authors.initials: L
authors.lastname: Capece
authors.affiliation: Dipartimento di Fisica e Scienze della Terra, Universitá degli Studi di Parma, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Pennacchietti
authors.affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina; IQUIBICEN-CONICET, Argentina.
authors.forename: Diana E
authors.initials: DE
authors.lastname: Wetzler
authors.affiliation: Dipartimento di Farmacia, Universitá degli Studi di Parma, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Bruno
authors.affiliation: Dipartimento di Fisica e Scienze della Terra, Universitá degli Studi di Parma, Italy.
authors.forename: Stefania
authors.initials: S
authors.lastname: Abbruzzetti
authors.affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina.
authors.forename: Lucía
authors.initials: L
authors.lastname: Chisari
authors.affiliation: Departament de Fisicoquímica and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Avda. Prat de la Riba 171, 08921 Santa Coloma de Gramenet, Spain.
authors.forename: F Javier
authors.initials: FJ
authors.lastname: Luque
authors.affiliation: Dipartimento di Fisica e Scienze della Terra, Universitá degli Studi di Parma, Italy.
authors.forename: Cristiano
authors.initials: C
authors.lastname: Viappiani
authors.affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina; INQUIMAE-CONICET, Argentina.
authors.forename: Marcelo A
authors.initials: MA
authors.lastname: Marti
authors.affiliation: Departamento de Química Inorgánica, Analítica y Química Física, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina; INQUIMAE-CONICET, Argentina. Electronic address: dario@qi.fcen.uba.ar.
authors.forename: Dario A
authors.initials: DA
authors.lastname: Estrin
authors.affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, Buenos Aires C1428EHA, Argentina; IQUIBICEN-CONICET, Argentina. Electronic address: anadra@qi.fcen.uba.ar.
authors.forename: Alejandro D
authors.initials: AD
authors.lastname: Nadra
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D005914:Globins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006418:Heme / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008024:Ligands; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009211:Myoglobin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000078787:Neuroglobin; D011485:Protein Binding; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry
chemical_list: D008024:Ligands; D009211:Myoglobin; D009419:Nerve Tissue Proteins; D000078787:Neuroglobin; D011993:Recombinant Fusion Proteins; D006418:Heme; D005914:Globins
keywords: Bis-histidyl coordination; Flash photolysis; Molecular dynamics; Myoglobin; Neuroglobin