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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

The 441.6-nm excited visible RR spectra of deoxyHb A (A), deoxy-rHb(αH87G) (B) and deoxy-rHb(βH92G) (C).The hemoglobin concentration was 200 μM (in heme) in a 0.05 M phosphate buffer, pH 7.0. In addition, rHb(αH87G) and rHb(βH92G) contained 10 mM imidazole. For rHb(αH87G), deconvoluted components, a νFe-His and a νFe-Im are indicated by a thin dotted red line and a brown solid line, respectively. For rHb(βH92G), deconvoluted components, a νFe-His (high), a νFe-His (low) and a νFe-Im are indicated by a thin dotted green, a solid blue line and a solid brown line, respectively.
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pone.0135080.g008: The 441.6-nm excited visible RR spectra of deoxyHb A (A), deoxy-rHb(αH87G) (B) and deoxy-rHb(βH92G) (C).The hemoglobin concentration was 200 μM (in heme) in a 0.05 M phosphate buffer, pH 7.0. In addition, rHb(αH87G) and rHb(βH92G) contained 10 mM imidazole. For rHb(αH87G), deconvoluted components, a νFe-His and a νFe-Im are indicated by a thin dotted red line and a brown solid line, respectively. For rHb(βH92G), deconvoluted components, a νFe-His (high), a νFe-His (low) and a νFe-Im are indicated by a thin dotted green, a solid blue line and a solid brown line, respectively.

Mentions: Fig 8 shows the 441.6 nm-excited resonance Raman spectra of Hb A (A), rHb(αH87G) (B) and rHb(βH92G) (C) in the deoxy form. Except for the bands around 220 cm-1 assignable to the Fe-His (νFe-His), or Fe-Im (νFe-Im) stretching and around 363 cm-1, the observed peak frequencies are almost the same among the three Hbs. The latter band is assigned to δ(CβCcCd)6,7, an in-plain bending mode of a propionate group [73]. This band appears at 363, 363, and 365 cm-1, for Hb A, rHb(αH87G) and rHb(βH92G), respectively. The δ(CβCcCd)6,7, mode shifts to a higher wavenumber in rHb(βH92G) than Hb A and rHb(αH87G), suggesting a subtle disorder of the tertiary structure around the propionate side-chain of the Fe-Im heme of the β subunits in rHb(βH92G).


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)

The 441.6-nm excited visible RR spectra of deoxyHb A (A), deoxy-rHb(αH87G) (B) and deoxy-rHb(βH92G) (C).The hemoglobin concentration was 200 μM (in heme) in a 0.05 M phosphate buffer, pH 7.0. In addition, rHb(αH87G) and rHb(βH92G) contained 10 mM imidazole. For rHb(αH87G), deconvoluted components, a νFe-His and a νFe-Im are indicated by a thin dotted red line and a brown solid line, respectively. For rHb(βH92G), deconvoluted components, a νFe-His (high), a νFe-His (low) and a νFe-Im are indicated by a thin dotted green, a solid blue line and a solid brown line, respectively.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0135080.g008: The 441.6-nm excited visible RR spectra of deoxyHb A (A), deoxy-rHb(αH87G) (B) and deoxy-rHb(βH92G) (C).The hemoglobin concentration was 200 μM (in heme) in a 0.05 M phosphate buffer, pH 7.0. In addition, rHb(αH87G) and rHb(βH92G) contained 10 mM imidazole. For rHb(αH87G), deconvoluted components, a νFe-His and a νFe-Im are indicated by a thin dotted red line and a brown solid line, respectively. For rHb(βH92G), deconvoluted components, a νFe-His (high), a νFe-His (low) and a νFe-Im are indicated by a thin dotted green, a solid blue line and a solid brown line, respectively.
Mentions: Fig 8 shows the 441.6 nm-excited resonance Raman spectra of Hb A (A), rHb(αH87G) (B) and rHb(βH92G) (C) in the deoxy form. Except for the bands around 220 cm-1 assignable to the Fe-His (νFe-His), or Fe-Im (νFe-Im) stretching and around 363 cm-1, the observed peak frequencies are almost the same among the three Hbs. The latter band is assigned to δ(CβCcCd)6,7, an in-plain bending mode of a propionate group [73]. This band appears at 363, 363, and 365 cm-1, for Hb A, rHb(αH87G) and rHb(βH92G), respectively. The δ(CβCcCd)6,7, mode shifts to a higher wavenumber in rHb(βH92G) than Hb A and rHb(αH87G), suggesting a subtle disorder of the tertiary structure around the propionate side-chain of the Fe-Im heme of the β subunits in rHb(βH92G).

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