Cooperative mechanism in the homodimeric myoglobin from Nassa mutabilis.
Oxygen binding and spectroscopic properties of the homodimeric myoglobin (Mb) from the prosobranchia sea snail Nassa mutabilis have been investigated. Oxygen equilibrium curves are pH-independent and cooperative with P50 = 5 +/- 1 mmHg and n approximately 1.5. Circular dichroism spectra of the oxygenated and deoxygenated form of N. mutabilis Mb are superimposable between 190 and 250 nm, suggesting a mechanism for cooperative ligand binding that does not involve changes in the alpha-helical content of the whole protein. The oxygen dissociation process is biphasic and pH-dependent, with different pKa values (=6.7 +/- 0.2 and 8.5 +/- 0.3) for the two phases. Moreover, the activation energy is essentially the same for both oxygen dissociation processes (Ea = 56.4 +/- 2.1 kJ/mol for the fast phase, and Ea = 53.8 +/- 1.9 kJ/mol for the slow phase), indicating that the rate difference for O2 dissociation between the diliganded and the monoliganded species is mostly dependent on a variation of the activation entropy. Ferrous nitrosylated N. mutabilis Mb shows, at alkaline and neutral pH, axial and rhombic X-band EPR signals, respectively, which display below pH 6 a three-hyperfine pattern typical of five-coordination. The results presented here suggest that in N.mutabilis Mb the kinetic control of cooperativity operates through a mechanism never observed before in other hemoproteins, which requires a ligand-linked large enhancement for the value of the oxygen association process in a molecule not undergoing changes in quaternary structure.
Coletta
M
M
Department of Experimental Medicine and Biochemical Sciences, University of Roma Tor Vergata, Via di Tor Vergata 135, 00133 Roma, Italy. coletta@seneca.ccd.utovrm.it
Ascenzi
P
P
Polizio
F
F
Smulevich
G
G
del Gaudio
R
R
Piscopo
M
M
Geraci
G
G
D000818:Animals; D049872:Bivalvia; D002942:Circular Dichroism; D006454:Hemoglobins / Q000737:chemistry; D008024:Ligands; D009211:Myoglobin / Q000737:chemistry* / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D012908:Snails
D006454:Hemoglobins; D008024:Ligands; D009211:Myoglobin; C083082:dimeric hemoglobin, Scapharca inaequivalvis; D010100:Oxygen