Hostname: page-component-78c5997874-4rdpn Total loading time: 0 Render date: 2024-11-02T18:54:24.986Z Has data issue: false hasContentIssue false

Oxygen binding by α(Fe2+)2β(Ni2+)2 hemoglobin crystals

Published online by Cambridge University Press:  01 April 2000

STEFANO BRUNO
Affiliation:
Institute of Biochemical Sciences, University of Parma, 43100 Parma, Italy
STEFANO BETTATI
Affiliation:
Institute of Biochemical Sciences, University of Parma, 43100 Parma, Italy
MICHELE MANFREDINI
Affiliation:
Institute of Biochemical Sciences, University of Parma, 43100 Parma, Italy
ANDREA MOZZARELLI
Affiliation:
Institute of Biochemical Sciences, University of Parma, 43100 Parma, Italy National Institute for the Physics of Matter, University of Parma, 43100 Parma, Italy
MARTINO BOLOGNESI
Affiliation:
Center of Advanced Biotechnology-IST and Department of Physics, National Institute for the Physics of Matter, University of Genova, 16132 Genova, Italy
DANIELA DERIU
Affiliation:
Center of Advanced Biotechnology-IST and Department of Physics, National Institute for the Physics of Matter, University of Genova, 16132 Genova, Italy
CAMILLO ROSANO
Affiliation:
Center of Advanced Biotechnology-IST and Department of Physics, National Institute for the Physics of Matter, University of Genova, 16132 Genova, Italy
ANTONIO TSUNESHIGE
Affiliation:
Department of Biochemistry and Biophysics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104
TAKASHI YONETANI
Affiliation:
Department of Biochemistry and Biophysics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104
ERIC R. HENRY
Affiliation:
Laboratory of Chemical Physics, NIDDK, NIH, Bethesda, Maryland 20892
Get access

Abstract

Oxygen binding by hemoglobin fixed in the T state either by crystallization or by encapsulation in silica gels is apparently noncooperative. However, cooperativity might be masked by different oxygen affinities of α and β subunits. Metal hybrid hemoglobins, where the noniron metal does not bind oxygen, provide the opportunity to determine the oxygen affinities of α and β hemes separately. Previous studies have characterized the oxygen binding by α(Ni2+)2 β(Fe2+)2 crystals. Here, we have determined the three-dimensional (3D) structure and oxygen binding of α(Fe2+)2 β(Ni2+)2 crystals grown from polyethylene glycol solutions. Polarized absorption spectra were recorded at different oxygen pressures with light polarized parallel either to the b or c crystal axis by single crystal microspectrophotometry. The oxygen pressures at 50% saturation (p50s) are 95 ± 3 and 87 ± 4 Torr along the b and c crystal axes, respectively, and the corresponding Hill coefficients are 0.96 ± 0.06 and 0.90 ± 0.03. Analysis of the binding curves, taking into account the different projections of the α hemes along the optical directions, indicates that the oxygen affinity of α1 hemes is 1.3-fold lower than α2 hemes. Inspection of the 3D structure suggests that this inequivalence may arise from packing interactions of the Hb tetramer within the monoclinic crystal lattice. A similar inequivalence was found for the β subunits of α(Ni2+)2 β(Fe2+)2 crystals. The average oxygen affinity of the α subunits (p50 = 91 Torr) is about 1.2-fold higher than the β subunits (p50 = 110 Torr). In the absence of cooperativity, this heterogeneity yields an oxygen binding curve of Hb A with a Hill coefficient of 0.999. Since the binding curves of Hb A crystals exhibit a Hill coefficient very close to unity, these findings indicate that oxygen binding by T-state hemoglobin is noncooperative, in keeping with the Monod, Wyman, and Changeux model.

Type
Research Article
Copyright
© 2000 The Protein Society

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)