Selective Attachment of Polyethylene Glycol to Hemerythrin for Potential Use in Blood Substitutes.

Mariann-Kinga Arkosi, Augustin C Mot, Iulia Lupan, Miruna Georgiana Ghinia Tegla, Radu Silaghi-Dumitrescu

The protein journal 2023 Aug

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Due to its ability to reversibly bind O2, alongside a relatively low redox reactivity and a limited cytotoxicity, the oxygen-carrying protein hemerythrin has been considered as an alternative to hemoglobin in preparing blood substitutes. In order to increase the hydrodynamic volume and lower antigenicity, two site-directed variants, H82C and K92C, were engineered that contained a single cysteine residue on the surface of each hemerythrin octamer for the specific attachment of polyethylene glycol (PEG). A sulfhydryl-reactive PEGylation reagent with a 51.9 Å spacer arm was used for selective cysteine derivatization. The mutants were characterized by UV-vis spectroscopy, size-exclusion chromatography, oxygen affinity, and autooxidation rate measurements. The H82C variant showed altered oligomeric behavior compared to the wild-type and was unstable in the met form. The PEGylated K92C variant is reasonably stable, displays an oxygen affinity similar to that of the wild-type, and shows an increased rate of autoxidation; the latter disadvantage may be counteracted by further chemical modifications. © 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.

Citation

Mariann-Kinga Arkosi, Augustin C Mot, Iulia Lupan, Miruna Georgiana Ghinia Tegla, Radu Silaghi-Dumitrescu. Selective Attachment of Polyethylene Glycol to Hemerythrin for Potential Use in Blood Substitutes. The protein journal. 2023 Aug;42(4):374-382