Histidine and not tyrosine is required for the Peroxide‐induced formation of haem to protein cross‐linked myoglobin

Abstract

SummaryPeroxide-induced oxidative modifications of haem proteins such as myoglobin and haemoglobin can lead to the formation of a covalent bond between the haem and globin. These haem to protein cross-linked forms of myoglobin and haemoglobin are cytotoxic and have been identified in pathological conditions in vivo. An understanding of the mechanism of haem to protein cross-link formation could provide important information on the mechanisms of the oxidative processes that lead to pathological complications associated with the formation of these altered myoglobins and haemoglobins. We have re-examined the mechanism of the formation of haem to protein cross-link to test the previously reported hypothesis that the haem forms a covalent bond to the protein via the tyrosine 103 residue (Catalano, C. E., Choe, Y. S., Ortiz de Montellano, P. R., J. Biol. Chem. 1989, 10534 -10541).Comparison of native horse myoglobin, recombinant sperm whale myoglobin and Tyr 103 ! Phe sperm whale mutant shows that, contrary to the previously proposed mechanism of haem to protein cross-link formation, the absence of tyrosine 103 has no impact on the formation of haem to protein cross-links. In contrast, we have found that engineered myoglobins that lack the distal histidine residue either cannot generate haem to protein cross-links or show greatly suppressed levels of modified protein. Moreover, addition of a distal histidine to myoglobin from Aplysia limacina, that naturally lacks this histidine, restores the haem protein's capacity to generate haem to protein cross-links. The distal histidine is, therefore, vital for the formation of haem to protein cross-link and we explore this outcome. IUBMB Life, 59: 477-489, 2007

Haptoglobin Binding Stabilizes Hemoglobin Ferryl Iron and the Globin Radical on Tyrosine β145

“…However, we show that this is not the case. Heme:protein cross-linking is a reaction catalyzed inside the heme pocket between the protonated ferryl and a nearby protein radical, probably the distal histidine (36). Consistent with the lack of effect on the ferryl pK, Hp does not alter the extent of this peroxide-induced cross-linking across a range of pH values.…”

“…The separate rate constants were calculated as described previously (37). The formation of covalent heme:protein cross-links (Hb-X) was determined by reversed phase HPLC as described previously (36).…”

Redox Reactions of Myoglobin

“…Cross-linked Mb is a green pigment, which may be due to disruption of the conjugation of one of the pyrrole rings. The distal histidine (His64) has been implicated as a key residue that facilitates formation of cross-linked Mb rather than Tyr103 (57).…”

The broad diversity of heme-protein cross-links: An overview

“…To further examine the mechanism for H 2 O 2 -induced crosslinking in Mb, Reeder et al [141] removed the Tyr103 (Y103F mutation) in sperm whale Mb (swMb) (Fig. 15, blue, PDB ID: 1JP6 [142]) and found it has almost no impact on the formation of crosslink, with an extend (25.2%) comparable to that of WT swMb (26.7%) at pH 5.0.…”