Chloride Control of the Mechanism of Human Serum Ceruloplasmin (Cp) Catalysis

Abstract

Unraveling the mechanism of ceruloplasmin (Cp) is fundamentally important towards understanding the pathogenesis of metal-mediated diseases and metal neurotoxicity. Here we report that Cl−, the most abundant anion in blood plasma, is a key component of Cp catalysis. Based on detailed spectroscopic analyses, Cl− preferentially interacts with the partially reduced trinuclear Cu cluster (TNC) in Cp under physiological conditions and shifts the electron equilibrium distribution among the two redox active type 1 (T1) Cu sites and the TNC. This shift in potential enables the intramolecular electron transfer (IET) from the T1 Cu to the native intermediate (NI) and accelerates the IET from the T1 Cu to the TNC, resulting in faster turnover in Cp catalysis.

Inhibition in multicopper oxidases: a critical review

“…DFT has been applied to the understanding the mechanism of Cl − activation of O 2 reduction in ceruloplasmin (section 3.2). 203 The calculations showed Cl − bridging between the T2 Cu and one T3 Cu (Fig. 12) and changing the ligand geometry around the T2 Cu, consistent with the authors' EPR measurements.…”

“…165,194 Solomon et al recently reported that Cl − binds preferentially to the partially reduced TNC of a human ceruloplasmin under physiological NaCl concentrations and pH. 203 They also noted a 10-fold increase in the reaction rate for o-dianisidine oxidation in the presence of 100 mM NaCl (at pH 7), confirming that Cl − can enhance the catalytic activity of ceruloplasmin under certain conditions.…”

Role of a Tyrosine Radical in Human Ceruloplasmin Catalysis

“… 11 Fully reduced Cp undergoes a similar pathway ( Figure S7 , bottom), and our previous studies showed that Cp requires a fully reduced TNC to achieve fast reaction with O 2 (∼10 6 M –1 s –1 ). 22 However, we have demonstrated in this work that partially reduced forms of Cp (3e – - or 4e – -reduced) are able to reduce O 2 by recruiting one extra electron from a nearby tyrosine residue. To achieve this rapid reaction, the first step likely involves a proton-coupled electron transfer (PCET) process from Tyr107 to the T2 Cu to generate the fully reduced TNC ( Figure S7 , top).…”

“…Recently we reported that chloride, the most abundant anion present in plasma (with a [Cl – ] ≈ 0.1 M), binds to the partially reduced TNC and shifts the electron equilibrium distribution between T1 Cu sites and the TNC in the Cp reduction process. 22 The Cl – -binding-induced reduction potential separation enables controlled formation of a series of partially reduced forms of Cp with well-defined electron distributions, as shown in Scheme 2 . The fully oxidized Cp is reduced stepwise following the order of decreasing reduction potentials.…”

“…The addition of one equivalent of reductant results in 50% formation of 2e – ). 22 The definition of these reduced states serves as an ideal entry point to dissect the reaction pathway of O 2 reduction by Cp. In this study we explored the O 2 reactivity of these systematically reduced Cp forms.…”