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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

Left: The CD spectral changes due to the quaternary and tertiary structure transition for Hb A.The spectra are the quaternary structure transition (A: pink spectrum) and tertiary structure transition (B: blue spectrum) expected for Hb A and the observed deoxy-minus-oxy difference spectra of Hb A (C: black spectrum). Right: Comparison of the deoxy-minus-oxy difference spectra of rHb(αH87G) and rHb(βH92G) with that of Hb A. The difference spectra are Hb A (black spectrum), rHb(αH87G) (orange spectrum) and rHb(βH92G) (green spectrum).
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pone.0135080.g010: Left: The CD spectral changes due to the quaternary and tertiary structure transition for Hb A.The spectra are the quaternary structure transition (A: pink spectrum) and tertiary structure transition (B: blue spectrum) expected for Hb A and the observed deoxy-minus-oxy difference spectra of Hb A (C: black spectrum). Right: Comparison of the deoxy-minus-oxy difference spectra of rHb(αH87G) and rHb(βH92G) with that of Hb A. The difference spectra are Hb A (black spectrum), rHb(αH87G) (orange spectrum) and rHb(βH92G) (green spectrum).

Mentions: The deoxy-minus-oxy difference CD spectrum of Hb A are illustrated by a black curve in the left panel of Fig 10, where the sum of the contributions from the four aromatic residues mentioned above to the quaternary structure transition is represented by a pink curve (A). The contributions from the tertiary structure changes to the CD spectra upon the change from deoxy (t) to oxy (r) state were estimated from the deoxy-minus-oxy differences of the isolated subunits [79] and superimposed on the same figure with a blue curve (B). It is stressed that a negative CD band is present around 294 nm for the t–r difference, at a longer wavelength than that for the T–R difference. Although the aromatic residues responsible for the negative t–r CD band have not been identified, Trp residues in both chains (Trpα14 and Trpβ15) would be involved.


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)

Left: The CD spectral changes due to the quaternary and tertiary structure transition for Hb A.The spectra are the quaternary structure transition (A: pink spectrum) and tertiary structure transition (B: blue spectrum) expected for Hb A and the observed deoxy-minus-oxy difference spectra of Hb A (C: black spectrum). Right: Comparison of the deoxy-minus-oxy difference spectra of rHb(αH87G) and rHb(βH92G) with that of Hb A. The difference spectra are Hb A (black spectrum), rHb(αH87G) (orange spectrum) and rHb(βH92G) (green spectrum).
© Copyright Policy
Related In: Results  -  Collection

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

pone.0135080.g010: Left: The CD spectral changes due to the quaternary and tertiary structure transition for Hb A.The spectra are the quaternary structure transition (A: pink spectrum) and tertiary structure transition (B: blue spectrum) expected for Hb A and the observed deoxy-minus-oxy difference spectra of Hb A (C: black spectrum). Right: Comparison of the deoxy-minus-oxy difference spectra of rHb(αH87G) and rHb(βH92G) with that of Hb A. The difference spectra are Hb A (black spectrum), rHb(αH87G) (orange spectrum) and rHb(βH92G) (green spectrum).
Mentions: The deoxy-minus-oxy difference CD spectrum of Hb A are illustrated by a black curve in the left panel of Fig 10, where the sum of the contributions from the four aromatic residues mentioned above to the quaternary structure transition is represented by a pink curve (A). The contributions from the tertiary structure changes to the CD spectra upon the change from deoxy (t) to oxy (r) state were estimated from the deoxy-minus-oxy differences of the isolated subunits [79] and superimposed on the same figure with a blue curve (B). It is stressed that a negative CD band is present around 294 nm for the t–r difference, at a longer wavelength than that for the T–R difference. Although the aromatic residues responsible for the negative t–r CD band have not been identified, Trp residues in both chains (Trpα14 and Trpβ15) would be involved.

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