The Protein Journal

, Volume 26, Issue 4, pp 257–263 | Cite as

Temperature Dependent Soret Spectral Band Shifts Accompany Human CN-Mesohemoglobin Assembly

  • Priyani  V. Fonseka
  • Gayathri Vasudevan
  • Lisa-Jo  Ann Clarizia
  • Melisenda  J. McDonald


The interaction between human apohemoglobin A and CN-Mesohemin, a monomeric non-native heme derivative, was probed by Soret spectrophotometric titrations in 0.05 M potassium phosphate buffer, pH 7 at varied temperatures. Hypsochromic shifts in the absorbance maxima were observed at all temperatures below 10°C. First derivative spectroscopy of CN-Mesohemin titrations was used to provide further evidence of a spectral shift upon CN-Mesohemoglobin assembly. Findings of Soret Spectral shifts demonstrate a preference for the α chain heme site by CN-Mesohemin indicative of semi-α-hemoglobin intermediate formation. CN-Mesohemin, a derivative with peripheral 2,4 ethyl groups, does not possess the extended conjugation seen for native CN-Protohemin with its 2,4 vinyl groups. Indeed, reduced polarity of CN-Mesohemin over that of CN-Protohemin resulted in distinct temperature dependencies. Molecular visualization and protein-ligand interaction analysis targeted a functionally diverse residue unique to the α-chain. Tyrosine-42 (a polar/non-polar amino acid) appeared to play a prominent role in the assembly process.


Apohemoglobin A CN-Mesohemin protein–ligand interactions Soret region 


  1. Ackers G. K., Holt J. M., Huang Y., Grinkova Y., Klinger A. L., Denisou I. (2000) Protein 4:23–43CrossRefGoogle Scholar
  2. Ascoli F., Fanelli M. R. R., Antonini E. (1981) Meth. Enzymol. 76:72–87PubMedGoogle Scholar
  3. Boffi A., Zamparelli C., Verzili D., Ilari A., Chiancone E. (1997) Archives of Biochem. Biophys. 340:43–51CrossRefGoogle Scholar
  4. Brown S. B. (1980) An Introduction to Spectroscopy for Biochemists. Academic Press, A Subsidiary of Harcourt Brace Jovanovich, Publishers, London New YorkGoogle Scholar
  5. Bunn H. F., Forget G. B. (1986) In: Dyson J. (ed) Hemoglobin: Molecular, Genetic and Clinical aspects. Saunders, Philadelphia, PA, pp 13–90Google Scholar
  6. Fischer H., Orth H. (1937) Die Chemie des pyrroles, Pyrrolfarbstoffe. II. Erste Halfte. Leipzig Akcadem, Verlagsgesellschaft, pp. 372Google Scholar
  7. Gibson Q. H. (1997) Archives of Biochem. Biophys. 339: 275–282CrossRefGoogle Scholar
  8. Gibson Q. H., Antonini E. (1963) J. Biol. Chem. 238: 1384–1388PubMedGoogle Scholar
  9. Ho C. (1992) Adv. Pro. Chem. 43: 153–312ADSCrossRefGoogle Scholar
  10. Ishimori K., Morishima I. (1988) Biochemistry 27: 4747–4753PubMedCrossRefGoogle Scholar
  11. Jennings T. M., McDonald M. J. (2002) Biochem. Biophys. Res. Comm. 293: 1354–1357PubMedCrossRefGoogle Scholar
  12. Joshi A. A., McDonald M. J. (1994) J. Biol. Chem.262: 5951–5956Google Scholar
  13. Leutzinger Y., Beychok S. (1981) Proc. Natl. Acad. Sci. USA 78: 780–781PubMedCrossRefADSGoogle Scholar
  14. Park and Tame: To be published (PDB ID# 1IRD)Google Scholar
  15. Perkampus H.H. (1992) UV-VIS Spectroscopy and its Applications. Springer Laboratory, Berlin Heidelberg, pp 88–94Google Scholar
  16. Perutz M. F., Wilkinson A. J., Paoli M., Dodson G. G. (1998) Annu. Rev. Biophys. Biomol Struct. 27: 1–34PubMedCrossRefGoogle Scholar
  17. Smith K. M. (1975) Porphyrins and Metalloporphyrins. Elsevier Scientific Publishing Company, Amsterdam-Oxford-New YorkGoogle Scholar
  18. Sobolev V., Sorokine A., Prilusky J., Abola E. E., Edelman M. (1999) Bioinformatics 15: 327–332PubMedCrossRefGoogle Scholar
  19. Stamatoyannopoulos G., Nienhuis A. W., Leader P., Majerus P. W. (1987) In: Dyson J. (ed) The Molecular Basis of Blood Disease, W.B. Saunders, Philadelphia PA, pp. 28–65Google Scholar
  20. Vasudevan G., McDonald M. J. (1997) J. Biol. Chem. 272: 517–524PubMedCrossRefGoogle Scholar
  21. Vasudevan G., McDonald M. J. (2002) Curr. Prot. Pept. Sci. 3: 461–466CrossRefGoogle Scholar
  22. Vasudevan G., McDonald M. J. (2006) The Prot. J. 25: 45–56CrossRefGoogle Scholar
  23. Yamaguchi T., Pang J., Reddy K. S., Witkowska H. E., Surrey S., Adachi K. (1996) J. Biol. Chem. 271: 26677–26683PubMedCrossRefGoogle Scholar
  24. Zamperilli C., Boffi A., Verzili D., Chiancone E., Rousseau D.L., Takahashi S., Sugita Y., Yoneyama Y. (1971) J. Biol. Chem. 246:389–394Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2006

Authors and Affiliations

  • Priyani  V. Fonseka
    • 1
    • 2
  • Gayathri Vasudevan
    • 1
    • 3
  • Lisa-Jo  Ann Clarizia
    • 1
  • Melisenda  J. McDonald
    • 1
  1. 1.Department of ChemistryUniversity of MassachusettsLowellUSA
  2. 2.Columbus Children’s Hospital Research InstituteColumbusUSA
  3. 3.Department of ChemistryWellesley CollegeWellesleyUSA

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