Electrochemical analysis of gold nanoparticles multifunctionalised withCytochrome c and a zinc Porphyrin

Cytochrome c (Cyt c), known for its functional redox capabilities, plays a pivotal role in biological

processes such as the electron transport chain and apoptosis. However, understanding how different

conjugation strategies impact its structural and redox characteristics is limited. To fill this gap, we

investigated the effects of conjugating Cyt c and a zinc(II) porphyrin (Zn Porph) to gold nanoparticles

(AuNPs). We used circular dichroism (CD) spectroscopy to detect structural conformational changes

in Cyt c upon conjugation and time-of-flight secondary ion mass spectrometry (TOF-SIMS) to

identify protein orientation. Cyt c was predicted to have different orientations depending on the size

of AuNPs and methods used to conjugate the protein, it was hypothesised that the orientation of Cyt

https://doi.org/10.26434/chemrxiv-2023-rsrwv ORCID: https://orcid.org/0000-0002-4872-8928 Content not peer-reviewed by ChemRxiv. License: CC BY 4.0

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c may influence the redox properties of the protein. The electrochemical properties of Cyt c were

assessed using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). We used DPVbased

to determine the heterogeneous rate constant (k0). The results show a lower k0 for conjugated

Cyt c than free Cyt c, likely due to structural changes in the protein. The spatial orientation of Cyt c

had minimal influence on k0, while ligand density and AuNP size had an effect. The k0 value of Zn

Porph did not decrease on conjugation. Despite these changes, Cyt c and Zn Porph maintained their

electrochemical capabilities after conjugation.