Manganese and Cobalt in the Nonheme-Metal-Binding Site of a Biosynthetic Model of Heme-Copper Oxidase Superfamily Confer Oxidase Activity through Redox-Inactive Mechanism

Abstract

The presence of nonheme metal, such as copper and iron, in the heme-copper oxidase (HCO) superfamily is critical to the enzymatic activity of reducing O2 to H2O, but the exact mechanism the nonheme metal ion uses to confer and fine-tune the activity remains to be understood. We report that manganese and cobalt can bind to the same nonheme site and confer HCO activity in a heme-nonheme biosynthetic model in myoglobin. While the initial rates of O2 reduction by the Mn, Fe and Co derivatives are similar, the percentage of reaction active species formation are 7%, 4% and 1% and the total turnovers are 5.1 ± 1.1, 13.4 ± 0.7, and 82.5 ± 2.5, respectively. These results correlate with the trends of nonheme metal-binding dissociation constants (35 μM, 22 μM and 9 μM) closely, suggesting that tighter metal binding can prevent ROS release from the active site, lessen damage to the protein, and produce higher total turnover numbers. Detailed spectroscopic, electrochemical, and computational studies found no evidence of redox cycling of manganese or cobalt in the enzymatic reactions, and suggest that structural and electronic effects related to the presence of different nonheme metals lead to observed differences in reactivity. This study of the roles of nonheme metal ions beyond the Cu and Fe found in native enzymes has provided deeper insights into nature’s choice of metal ion, and reaction mechanism, and allows for finer control of the enzymatic activity, which is a basis for design of efficient catalysts for oxygen reduction reaction for fuel cells.

Circular Polarized Light Activated Chiral Satellite Nanoprobes for the Imaging and Analysis of Multiple Metal Ions in Living Cells

“…When the level of metal ions deviates from the normal range, disease, such as Alzheimer's and Parkinson's disease, and cancer cell proliferation, often occurs . As cofactors, metal ions participate in cellular behavior processes, especially zinc, magnesium, and copper, which participate in a series of physiological and cellular behaviors . Even though a series of chemical probes using either small fluorescent organic molecules or proteins have been reported for metal ion detection, the large number of different metal ions in living systems calls for a more generalizable method for imaging and quantifying as many metal ions as possible in living systems .…”

Synthetic Fe/Cu Complexes: Toward Understanding Heme-Copper Oxidase Structure and Function

“…Recently, the insertion of other first row transition metals into the NOR model Fe B Mb template uncovered further important details pertaining to the role of the nonheme metal in the O 2 -reduction reaction. 1098 Both cobalt and manganese are known to be active in metalloenzyme redox chemistry and importantly can bind in the nonheme site of Fe B Mb (UV-vis and crystallographic evidence), thereby making Mn II -Fe B Mb and Co II -Fe B Mb good candidates for a systematic study with the previously described Fe II -Fe B Mb (see just above). The binding affinity of these metals in the engineered nonheme site follows the Irving-Williams series trend (Mn II < Fe II < Co II ) and interestingly also correlates well with total turnover numbers and selectivity (Figure 141).…”

“…1093 It was also ruled out using spectroelectrochemical methods that the identity of the nonheme metal may be regulating the O 2 chemistry by impacting the heme-iron reduction potential, given the small shifts in E°' heme regardless of the nonheme metal and the lack of correlation to the O 2 -reduction activity of the models. 1093,1098,1099 To further understand the reason for the differences in selectivity across nonheme metal ion identities, the authors looked beyond the impact of tight metal binding which is undoubtedly important for stable, continuous turnover. The two factors which stand out with respect to the most selective model, Co II -Fe B Mb, are that (i) this variant forms the heme-oxy intermediate the slowest, and (ii) the Co II ion is the smallest and most Lewis acidic of the three ions employed in this study.…”

“…(C) Tabulated parameters including metal dissociation constants, initial rates of O 2 -consumption, total turnover numbers, and percentage of ROS formation for the four variants showing the trends described in the text. Adapted from ref 1098. Copyright 2017 American Chemical Society.…”

Circular Polarized Light Activated Chiral Satellite Nanoprobes for the Imaging and Analysis of Multiple Metal Ions in Living Cells

“…[3][4][5][6] As cofactors,m etal ions participate in cellular behavior processes,e specially zinc, magnesium, and copper, which participate in as eries of physiological and cellular behaviors. [7][8][9][10] Even though aseries of chemical probes using either small fluorescent organic molecules or proteins have been reported for metal ion detection, [8,[11][12][13] the large number of different metal ions in living systems calls for am ore generalizable method for imaging and quantifying as many metal ions as possible in living systems. [14] To meet this challenge,t he simultaneous detection and bioimaging of multiple metal ions in live cells is anecessary research direction.…”