Molecular pathology of dityrosine cross-links in proteins: Structural and functional analysis of four proteins

  • Dorairajan Balasubramanian
  • Ritu Kanwar
Part of the Developments in Molecular and Cellular Biochemistry book series (DMCB, volume 37)


The dityrosine bond (DT) is an oxidative covalent cross-link between two tyrosines. DT cross-linking is increasingly identified as a marker of oxidative stress, aging and disease, and has been detected in diverse pathologies. While DT cross-linked proteins have been documented, the consequences of the DT link on the structure and function of the so modified proteins are yet to be understood. With this in view, we have studied the properties of intermolecular DT-dimers of four proteins of diverse functions, namely the enzyme ribonuclease A, the signal protein calmodulin, and the eye lens proteins alpha-and gamma Bcrystallins. We find that DT is formed through radical reactions and type I photosensitization (including OH, O2 - and 00N0-), but not by 1O2and NO2 - (which modify his, trp and met more readily). Tyr residues on the surface of the protein make DT bonds (intra-and intermolecular) most readily and preferentially. The conformation of each of these DT-dimers, monitored by spectroscopy, is seen not to be significantly altered in comparison to that of the parent monomer, but the structural stability of the DT cross-linked molecule is lower than that of the parent native monomer. The DT-dimer is denatured at a lower temperature, and at lower concentrations of urea or guanidinium chloride. The effect of DT-cross-linking on the biological activities of these proteins was next studied. The enzymatic activity of the DT-dimer of ribonuclease A is not lost but lowered. DT-dimerization of lens alpha-crystallin did not significantly affect the chaperone-like ability; it inhibits the self-aggregation and precipitation of target proteins just as well as the parent, unmodified alpha-crystallin does. DT-dimerization of gamma B-crystallin is however seen to lead to more ready aggregation and precipitation, a point of interest in cataract. In the case of calmodulin, we could generate both intermolecular and intramolecular DT cross-linking, and study both the DT-dimer and DT-monomer. The DT-dimer binds smooth muscle light chain kinase and also Ca2+, but less efficiently and over a broad concentration range than the native monomer. The intramolecular DT-monomer is weaker in all these respects, presumably since it is structurally more constrained. These results suggest that DT cross-linking of globular proteins weakens their structural stability and compromises (though does not abolish) their biological activity, both of which are pathologically relevant. The intramolecular DT cross-link would appear to lead to more severe structural and functional consequences. (Mol Cell Biochem 234/235: 27–38, 2002)

Key words

dityrosine cross-link oxidative damage pathological marker chaperone-like activity cataract 


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Copyright information

© Springer Science+Business Media New York 2002

Authors and Affiliations

  • Dorairajan Balasubramanian
    • 1
  • Ritu Kanwar
    • 1
  1. 1.Hyderabad Eye Research FoundationL.V. Prasad Eye InstituteHyderabadIndia

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