Studies on the Properties of Fish Hemoglobins

Abstract

The ligand-binding kinetics of the isolated component from trout (Salmo irideus) blood has been extensively investigated ; it appears to be fully consistent with their equilibrium behaviour, described previously. At alkaline pH (z 7 4 , in a region where the Root effect is not operative, comparison of the kinetics of the various components (Hb trout I, 11 and IV) shows considerable similarities between them. For Hb trout I (and 11) the time course of the reaction with both 0, and CO is practically pH independent. On the other hand Hb trout IV, characterized by a Root effect, displays a strong dependence on pH of the kinetics of ligand binding. Thus in the pH region where the Root effect becomes operative there is a large change of the character, as well as of the speed, of the kinetics with both 0, and CO.The Root effect characteristic of Hb trout IV appears to originate, from a kinetic point of view, in a pH-dependent change of both the "on" and "off" constants, although a t least for 0, the effect of pH on the ''off' kinetic constants is much more marked.

Properties of trout HbI in water and ligand linked binding of Na+

“…The time course of CO recombination depends on the fractional photodissociation [lo] and as the flash intensity is decreased the presence of a quickly reacting component is easily detectable. These experiments in the absence ofbuffer show that all the main features of trout Hb I kinetics are maintained, since the overall results are very similar to those reported for the same protein in buffered solutions [2,8,10]. This implies that the kinetic contribution to cooperativity is an intrinsic property of TIME (msecj TIME (tIWeC) the molecule and is not determined to any substantial extent by heterotropic effects, at least for CO.…”

“…A crystal of sodium dithionite was added to ensure complete removal of oxygen; the maximum drop in pH (6.8) observed with the unbuffered solution has no effect on the time course of CO binding (see also 121). Analysis of the time course of binding observed under conditions of complete photolysis shows that the reaction is autocatalytic both in water and in buffer (see also [2]). In no case the full photolysis data show the presence of quickly reacting components, indicating that at PM levels trout Hb I in water is fully tetrameric even in the liganded form (similarly to what clearly demonstrated in buffered solution) [2].…”

“…Analysis of the time course of binding observed under conditions of complete photolysis shows that the reaction is autocatalytic both in water and in buffer (see also [2]). In no case the full photolysis data show the presence of quickly reacting components, indicating that at PM levels trout Hb I in water is fully tetrameric even in the liganded form (similarly to what clearly demonstrated in buffered solution) [2]. The shape of the time course is very similar for the two conditions ( fig.3).…”

Studies on the Properties of Fish Hemoglobins Molecular Properties and Interaction with Third Components of the Isolated Hemoglobins from Trout (Salmo irideus)

“…7 and accompanying paper The structural and functional properties of Hb trout I differ very markedly from those characteristic of human Hb: a ) A t neutral pH dissociation of the tetrarner into subunits is hardly measurable. To the very high stability of the tetramer may be related: (i) the absence of hybrid formation (see above) ; (ii) the difficulty of digesting Hb trout I in its native state with carboxypeptidase A or B [22] ; (iii) the absence of a quickly reacting photoproduct upon complete photodissociation (see accompanying paper [19]). b) The shape of the far ultraviolet CD spectrum is also substantially different, in the region around 222 nm, from that of both human hemoglobin and Hb trout IV.…”

“…The results for 0, are less clear cut, also in view of the fact that by flash photolysis of oxyhemoglobin trout IV, only partial (z 50°/,) photodissociation is achieved. For presentation and discussion of the kinetics of ligand binding by trout hemoglobin components, see following paper [19].…”