Probing the conformation of hemoglobin Presbyterian in the R-state

Seetharama A. Acharya, Ashok Malavalli, Eric Peterson, Philip D. Sun, Chien Ho, Muthuchidambaram Prabhakaran, Arthur Arnone, Belur N. Manjula, Joel M. Friedman

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

The influence of allosteric effectors on the R-state (liganded) conformation of Tg-HbP (human hemoglobin Presbyterian expressed in transgenic pig) has been probed using a number of biophysical techniques, and the results have been compared with that of liganded of HbA (human normal adult hemoglobin) to gain insight into the molecular basis of Asn-108(β)->Lys mutation-induced low-oxygen affinity of Hb. The nuclear magnetic resonance studies of Tg-HbP revealed that the conformation of the α 1β1, and α1β2 interfaces of the protein in the deoxy state are indistinguishable from that of deoxy HbA, whereas the conformation of the microenvironment of His-103(α) of Tg-HbP, a residue of the α1β1 interface, is distinct from that of HbA in the R-state. In addition, the Presbyterian mutation also influences the structure of oxy Hb in other regions of the molecule. First, it facilitates the generation of deoxy (T)-state marker at 14. 2 ppm (from 2,2-dimethyl-s-silapentane-5-sulfonate) on the interaction of oxy Hb with inositol hexa-phosphate without changing the ligation state. Second, it increases the geminate yield of the 10 ns photoproduct of CO-Hb. Third, it enhances the propensity of phosphate to increase the geminate yield. Fourth, it potentiates the ability of phosphate to induce deoxy-like features at the heme environment in the R-state. Fifth, it induces T-state-like signatures at the switch and hinge regions of the α1β2 interface. Finally, molecular modeling studies have indicated an increased affinity for the four anion binding sites mapped in the midcentral cavity of Hb caused by the presence of Lys-108(β). In short, Lys-108(β) in HbP induces a propensity for oxy Hb to access T-like conformational features in different regions of the oxy Hb molecule and also enhances the T-like signatures in the oxy state on interaction with allosteric effectors without changing its ligation. Interestingly, the intrinsic T-like conformational features of the R-state of HbP, in addition to those induced by the addition of allosteric effectors to liganded HbP, appear to be reminiscent of features of the B-state conformation of Hb found in rHb 1.1 (recombinant hemoglobin). We propose that the lowered oxygen affinity of Tg-HbP in the presence of allosteric effectors is a consequence of an altered R-state conformation of Hb, which reflects the facilitation of switching the R-state of HbP to the T-state compared with the normal R-state of HbA, thereby reducing HbA's affinity to oxygen.

Original languageEnglish (US)
Pages (from-to)221-230
Number of pages10
JournalJournal of Protein Chemistry
Volume22
Issue number3
DOIs
StatePublished - Apr 2003

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Hemoglobin
Conformations
Oxygen
Ligation
Hemoglobins
Phosphates
Mutation
Inositol Phosphates
Carbon Monoxide
Heme
Anions
Magnetic Resonance Spectroscopy
Swine
Binding Sites
Molecules
Molecular modeling
Inositol
Binding sites
Hinges
Negative ions

ASJC Scopus subject areas

  • Biochemistry

Cite this

Acharya, S. A., Malavalli, A., Peterson, E., Sun, P. D., Ho, C., Prabhakaran, M., ... Friedman, J. M. (2003). Probing the conformation of hemoglobin Presbyterian in the R-state. Journal of Protein Chemistry, 22(3), 221-230. https://doi.org/10.1023/A:1025080801951

Probing the conformation of hemoglobin Presbyterian in the R-state. / Acharya, Seetharama A.; Malavalli, Ashok; Peterson, Eric; Sun, Philip D.; Ho, Chien; Prabhakaran, Muthuchidambaram; Arnone, Arthur; Manjula, Belur N.; Friedman, Joel M.

In: Journal of Protein Chemistry, Vol. 22, No. 3, 04.2003, p. 221-230.

Research output: Contribution to journalArticle

Acharya, SA, Malavalli, A, Peterson, E, Sun, PD, Ho, C, Prabhakaran, M, Arnone, A, Manjula, BN & Friedman, JM 2003, 'Probing the conformation of hemoglobin Presbyterian in the R-state', Journal of Protein Chemistry, vol. 22, no. 3, pp. 221-230. https://doi.org/10.1023/A:1025080801951
Acharya SA, Malavalli A, Peterson E, Sun PD, Ho C, Prabhakaran M et al. Probing the conformation of hemoglobin Presbyterian in the R-state. Journal of Protein Chemistry. 2003 Apr;22(3):221-230. https://doi.org/10.1023/A:1025080801951
Acharya, Seetharama A. ; Malavalli, Ashok ; Peterson, Eric ; Sun, Philip D. ; Ho, Chien ; Prabhakaran, Muthuchidambaram ; Arnone, Arthur ; Manjula, Belur N. ; Friedman, Joel M. / Probing the conformation of hemoglobin Presbyterian in the R-state. In: Journal of Protein Chemistry. 2003 ; Vol. 22, No. 3. pp. 221-230.
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N2 - The influence of allosteric effectors on the R-state (liganded) conformation of Tg-HbP (human hemoglobin Presbyterian expressed in transgenic pig) has been probed using a number of biophysical techniques, and the results have been compared with that of liganded of HbA (human normal adult hemoglobin) to gain insight into the molecular basis of Asn-108(β)->Lys mutation-induced low-oxygen affinity of Hb. The nuclear magnetic resonance studies of Tg-HbP revealed that the conformation of the α 1β1, and α1β2 interfaces of the protein in the deoxy state are indistinguishable from that of deoxy HbA, whereas the conformation of the microenvironment of His-103(α) of Tg-HbP, a residue of the α1β1 interface, is distinct from that of HbA in the R-state. In addition, the Presbyterian mutation also influences the structure of oxy Hb in other regions of the molecule. First, it facilitates the generation of deoxy (T)-state marker at 14. 2 ppm (from 2,2-dimethyl-s-silapentane-5-sulfonate) on the interaction of oxy Hb with inositol hexa-phosphate without changing the ligation state. Second, it increases the geminate yield of the 10 ns photoproduct of CO-Hb. Third, it enhances the propensity of phosphate to increase the geminate yield. Fourth, it potentiates the ability of phosphate to induce deoxy-like features at the heme environment in the R-state. Fifth, it induces T-state-like signatures at the switch and hinge regions of the α1β2 interface. Finally, molecular modeling studies have indicated an increased affinity for the four anion binding sites mapped in the midcentral cavity of Hb caused by the presence of Lys-108(β). In short, Lys-108(β) in HbP induces a propensity for oxy Hb to access T-like conformational features in different regions of the oxy Hb molecule and also enhances the T-like signatures in the oxy state on interaction with allosteric effectors without changing its ligation. Interestingly, the intrinsic T-like conformational features of the R-state of HbP, in addition to those induced by the addition of allosteric effectors to liganded HbP, appear to be reminiscent of features of the B-state conformation of Hb found in rHb 1.1 (recombinant hemoglobin). We propose that the lowered oxygen affinity of Tg-HbP in the presence of allosteric effectors is a consequence of an altered R-state conformation of Hb, which reflects the facilitation of switching the R-state of HbP to the T-state compared with the normal R-state of HbA, thereby reducing HbA's affinity to oxygen.

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