Spectroscopic Characterization of a Green Copper Site in a Single-Domain Cupredoxin

Abstract

Cupredoxins are widespread copper-binding proteins, mainly involved in electron transfer pathways. They display a typical rigid greek key motif consisting of an eight stranded β-sandwich. A fascinating feature of cupredoxins is the natural diversity of their copper center geometry. These geometry variations give rise to drastic changes in their color, such as blue, green, red or purple. Based on several spectroscopic and structural analyses, a connection between the geometry of their copper-binding site and their color has been proposed. However, little is known about the relationship between such diversity of copper center geometry in cupredoxins and possible implications for function. This has been difficult to assess, as only a few naturally occurring green and red copper sites have been described so far. We report herein the spectrocopic characterization of a novel kind of single domain cupredoxin of green color, involved in a respiratory pathway of the acidophilic organism Acidithiobacillus ferrooxidans. Biochemical and spectroscopic characterization coupled to bioinformatics analysis reveal the existence of some unusual features for this novel member of the green cupredoxin sub-family. This protein has the highest redox potential reported to date for a green-type cupredoxin. It has a constrained green copper site insensitive to pH or temperature variations. It is a green-type cupredoxin found for the first time in a respiratory pathway. These unique properties might be explained by a region of unknown function never found in other cupredoxins, and by an unusual length of the loop between the second and the fourth copper ligands. These discoveries will impact our knowledge on non-engineered green copper sites, whose involvement in respiratory chains seems more widespread than initially thought.

Biohydrogen for a New Generation of H2/O2 Biofuel Cells: A Sustainable Energy Perspective

“…[333] Some bacterial metabolisms in the biodiversity pool are known to involve high-potential electron-transfer chains. [334,335] Identification of high redox potential CcOs would be an original way to address the issue of low oxygen availability in H 2 /O 2 EBFCs. This research is also a good illustration of how more studies into applications will increase our knowledge of enzyme electrochemistry, and help in the understanding of fundamental issues such as enzymatic reactions in complex electron-transfer chains.…”

Cytoplasmic CopZ-Like Protein and Periplasmic Rusticyanin and AcoP Proteins as Possible Copper Resistance Determinants in Acidithiobacillus ferrooxidans ATCC 23270

“…This protein was later proposed to be part of the iron-oxidizing/O 2 -reducing system in A. ferrooxidans (26,30). This periplasmic protein, now named AcoP, was recently characterized and also contains a cupredoxintype copper-binding site that binds one Cu atom per molecule and likewise forms part of the rus operon and the iron respiratory system (31). Furthermore, the levels of AcoP also increased in sulfur-grown cells in the presence of copper (29).…”

Microbial copper resistance: importance in biohydrometallurgy

“…This protein was later proposed to be part of the iron-oxidizing/O 2 -reducing system in A. ferrooxidans although its role has not been defined so far (Castelle et al, 2008(Castelle et al, , 2010. This periplasmic protein, now named AcoP, has recently been characterized and it also contains a cupredoxin type copper-binding site that binds one Cu atom per molecule and likewise forms part of the rus operon and iron respiratory system (Roger et al, 2014). Furthermore, the levels of AcoP also increased in sulfur-grown cells in the presence of copper (Alm arcegui et al, 2014a).…”

“…Furthermore, the levels of AcoP also increased in sulfur-grown cells in the presence of copper (Alm arcegui et al, 2014a). Rus and AcoP have conserved copper-binding sites (HXCX 2-4 HX 4 M and HXCX 4-8 HX 4 M respectively) (Roger et al, 2014) such as those described in CopC, a cupredoxin-like copper-binding protein from P. syringae involved in copper homeostasis (Arnesano et al, 2002).…”