Structural-functional characterization of the cathodic haemoglobin of the conger eel Conger conger: molecular modelling study of an additional phosphate-binding site.
The protein sequence data for the alpha- and beta-chains have been deposited in the SWISS-PROT and TrEMBL protein knowledgebase under the accession numbers P83479 and P83478 respectively. The Conger conger (conger eel) haemoglobin (Hb) system is made of three components, one of which, the so-called cathodic Hb, representing approx. 20% of the total pigment, has been purified and characterized from both a structural and functional point of view. Stripped Hb showed a reverse Bohr effect, high oxygen affinity and slightly low cooperativity in the absence of any effector. Addition of saturating GTP strongly influences the pH dependence of the oxygen affinity, since the reverse Bohr effect, observed under stripped conditions, is converted into a small normal Bohr effect. A further investigation of the GTP effect on oxygen affinity, carried out by fitting its titration curve, demonstrated the presence of two independent binding sites. Therefore, on the basis of the amino acid sequence of the alpha- and beta-chains, which have been determined, a computer modelling study has been performed. The data suggest that C. conger cathodic Hb may bind organic phosphates at two distinct binding sites located along the central cavity of the tetramer by hydrogen bonds and/or electrostatic interactions with amino acid residues of both chains, which have been identified. Among these residues, the two Lys-alpha(G6) (where the letter refers to the haemoglobin helix and the number to the amino acid position in the helix) appear to have a key role in the GTP movement from the external binding region to the internal central cavity of the tetrameric molecule.
Pellegrini
Mariagiuseppina
M
Department of Sciences Applied to Biosystems, University of Cagliari, Cittadella Universitaria, I-09042 Monserrato (CA), Italy. pelleg@unica.it
Giardina
Bruno
B
Verde
Cinzia
C
Carratore
Vito
V
Olianas
Alessandra
A
Sollai
Luigi
L
Sanna
Maria T
MT
Castagnola
Massimo
M
di Prisco
Guido
G
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001665:Binding Sites; D004524:Eels / Q000097:blood*; D006160:Guanosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010100:Oxygen / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D000596:Amino Acids; D006454:Hemoglobins; D006160:Guanosine Triphosphate; D010100:Oxygen