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An Origin of Cooperative Oxygen Binding of Human Adult Hemoglobin: Different Roles of the α and β Subunits in the α2β2 Tetramer.

Nagatomo S, Nagai Y, Aki Y, Sakurai H, Imai K, Mizusawa N, Ogura T, Kitagawa T, Nagai M - PLoS ONE (2015)

Bottom Line: Resonance Raman, 1H NMR, and near-UV circular dichroism measurements revealed that the quaternary structure change did not occur upon O2-binding to rHb(αH87G), but it did partially occur with O2-binding to rHb(βH92G).The quaternary structure of rHb(αH87G) appears to be frozen in T while its tertiary structure is changeable.Thus, the absence of the Fe-His bond in the α subunit inhibits the T to R quaternary structure change upon O2-binding, but its absence in the β subunit simply enhances the O2-affinity of α subunit.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Tsukuba, Tsukuba, Ibaraki, Japan.

ABSTRACT
Human hemoglobin (Hb), which is an α2β2 tetramer and binds four O2 molecules, changes its O2-affinity from low to high as an increase of bound O2, that is characterized by 'cooperativity'. This property is indispensable for its function of O2 transfer from a lung to tissues and is accounted for in terms of T/R quaternary structure change, assuming the presence of a strain on the Fe-histidine (His) bond in the T state caused by the formation of hydrogen bonds at the subunit interfaces. However, the difference between the α and β subunits has been neglected. To investigate the different roles of the Fe-His(F8) bonds in the α and β subunits, we investigated cavity mutant Hbs in which the Fe-His(F8) in either α or β subunits was replaced by Fe-imidazole and F8-glycine. Thus, in cavity mutant Hbs, the movement of Fe upon O2-binding is detached from the movement of the F-helix, which is supposed to play a role of communication. Recombinant Hb (rHb)(αH87G), in which only the Fe-His in the α subunits is replaced by Fe-imidazole, showed a biphasic O2-binding with no cooperativity, indicating the coexistence of two independent hemes with different O2-affinities. In contrast, rHb(βH92G), in which only the Fe-His in the β subunits is replaced by Fe-imidazole, gave a simple high-affinity O2-binding curve with no cooperativity. Resonance Raman, 1H NMR, and near-UV circular dichroism measurements revealed that the quaternary structure change did not occur upon O2-binding to rHb(αH87G), but it did partially occur with O2-binding to rHb(βH92G). The quaternary structure of rHb(αH87G) appears to be frozen in T while its tertiary structure is changeable. Thus, the absence of the Fe-His bond in the α subunit inhibits the T to R quaternary structure change upon O2-binding, but its absence in the β subunit simply enhances the O2-affinity of α subunit.

No MeSH data available.


Related in: MedlinePlus

Schematic presentation of the normal heme (left) and the cavity mutant heme (F8His→Gly) (right).The crystal structure of the cavity mutant Mb, rMb(H93G), determined in the presence of imidazole, revealed that an imidazole molecule is bonded to the heme iron on the proximal side, as shown here (Barrick, D. et al., Biochemistry1994, 33, 6546−6554). Atomic coordinates of 2DN2 (ref. 46) were used about description of ribbon model of F-helix.
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pone.0135080.g001: Schematic presentation of the normal heme (left) and the cavity mutant heme (F8His→Gly) (right).The crystal structure of the cavity mutant Mb, rMb(H93G), determined in the presence of imidazole, revealed that an imidazole molecule is bonded to the heme iron on the proximal side, as shown here (Barrick, D. et al., Biochemistry1994, 33, 6546−6554). Atomic coordinates of 2DN2 (ref. 46) were used about description of ribbon model of F-helix.

Mentions: Therefore, we planned to prepare cavity mutant Hbs, rHb(αH87G) and rHb(βH92G), which can bind four O2 molecules per one Hb molecule. The cavity mutant Hbs were first constructed by Ho’s group, and its ligand binding properties were examined using n-butylisocyanide and 1H NMR spectra [54]. As illustrated in Fig 1, the cavity mutant Hbs have similar properties to hybrid Hbs regarding the absence of the Fe-His bond in either the α or β subunit, for which a movement of the Fe-Im bond concomitant with ligand binding is not directly communicated to the F-helix [54]. In addition, the cavity mutant Hbs can take the fully ligand-bound form, because O2 or CO can bind to the imidazole-bound heme, and therefore, the effects of the interactions between bound ligand and the protein in the distal pocket are maintained.


An Origin of Cooperative Oxygen Binding of Human Adult Hemoglobin: Different Roles of the α and β Subunits in the α2β2 Tetramer.

Nagatomo S, Nagai Y, Aki Y, Sakurai H, Imai K, Mizusawa N, Ogura T, Kitagawa T, Nagai M - PLoS ONE (2015)

Schematic presentation of the normal heme (left) and the cavity mutant heme (F8His→Gly) (right).The crystal structure of the cavity mutant Mb, rMb(H93G), determined in the presence of imidazole, revealed that an imidazole molecule is bonded to the heme iron on the proximal side, as shown here (Barrick, D. et al., Biochemistry1994, 33, 6546−6554). Atomic coordinates of 2DN2 (ref. 46) were used about description of ribbon model of F-helix.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526547&req=5

pone.0135080.g001: Schematic presentation of the normal heme (left) and the cavity mutant heme (F8His→Gly) (right).The crystal structure of the cavity mutant Mb, rMb(H93G), determined in the presence of imidazole, revealed that an imidazole molecule is bonded to the heme iron on the proximal side, as shown here (Barrick, D. et al., Biochemistry1994, 33, 6546−6554). Atomic coordinates of 2DN2 (ref. 46) were used about description of ribbon model of F-helix.
Mentions: Therefore, we planned to prepare cavity mutant Hbs, rHb(αH87G) and rHb(βH92G), which can bind four O2 molecules per one Hb molecule. The cavity mutant Hbs were first constructed by Ho’s group, and its ligand binding properties were examined using n-butylisocyanide and 1H NMR spectra [54]. As illustrated in Fig 1, the cavity mutant Hbs have similar properties to hybrid Hbs regarding the absence of the Fe-His bond in either the α or β subunit, for which a movement of the Fe-Im bond concomitant with ligand binding is not directly communicated to the F-helix [54]. In addition, the cavity mutant Hbs can take the fully ligand-bound form, because O2 or CO can bind to the imidazole-bound heme, and therefore, the effects of the interactions between bound ligand and the protein in the distal pocket are maintained.

Bottom Line: Resonance Raman, 1H NMR, and near-UV circular dichroism measurements revealed that the quaternary structure change did not occur upon O2-binding to rHb(αH87G), but it did partially occur with O2-binding to rHb(βH92G).The quaternary structure of rHb(αH87G) appears to be frozen in T while its tertiary structure is changeable.Thus, the absence of the Fe-His bond in the α subunit inhibits the T to R quaternary structure change upon O2-binding, but its absence in the β subunit simply enhances the O2-affinity of α subunit.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Tsukuba, Tsukuba, Ibaraki, Japan.

ABSTRACT
Human hemoglobin (Hb), which is an α2β2 tetramer and binds four O2 molecules, changes its O2-affinity from low to high as an increase of bound O2, that is characterized by 'cooperativity'. This property is indispensable for its function of O2 transfer from a lung to tissues and is accounted for in terms of T/R quaternary structure change, assuming the presence of a strain on the Fe-histidine (His) bond in the T state caused by the formation of hydrogen bonds at the subunit interfaces. However, the difference between the α and β subunits has been neglected. To investigate the different roles of the Fe-His(F8) bonds in the α and β subunits, we investigated cavity mutant Hbs in which the Fe-His(F8) in either α or β subunits was replaced by Fe-imidazole and F8-glycine. Thus, in cavity mutant Hbs, the movement of Fe upon O2-binding is detached from the movement of the F-helix, which is supposed to play a role of communication. Recombinant Hb (rHb)(αH87G), in which only the Fe-His in the α subunits is replaced by Fe-imidazole, showed a biphasic O2-binding with no cooperativity, indicating the coexistence of two independent hemes with different O2-affinities. In contrast, rHb(βH92G), in which only the Fe-His in the β subunits is replaced by Fe-imidazole, gave a simple high-affinity O2-binding curve with no cooperativity. Resonance Raman, 1H NMR, and near-UV circular dichroism measurements revealed that the quaternary structure change did not occur upon O2-binding to rHb(αH87G), but it did partially occur with O2-binding to rHb(βH92G). The quaternary structure of rHb(αH87G) appears to be frozen in T while its tertiary structure is changeable. Thus, the absence of the Fe-His bond in the α subunit inhibits the T to R quaternary structure change upon O2-binding, but its absence in the β subunit simply enhances the O2-affinity of α subunit.

No MeSH data available.


Related in: MedlinePlus