Engineering a bifunctional copper site in the cupredoxin fold by loop-directed mutagenesis

ESPINOZA-CARA, ANDRÉS; ZITARE, ULISES; ALVAREZ-PAGGI, DAMIÁN; KLINKE, SEBASTIÁN; OTERO, LISANDRO H.; MURGIDA, DANIEL H.; VILA, ALEJANDRO J.

Chemical Science

Royal Society of Chemistry

Año: 2018 vol. 9 p. 6692 - 6702

2041-6520

Coppersites in proteins are designed to perform either electron transfer orredox catalysis. Type 1 and Cu A sites are electron transfer hubs bound to a rigid protein fold that prevents binding of exogenous ligands and side reactions. Here we report the engineering of two Type 1 sites by loop-directed mutagenesis within a CuA scaffold with unique electronic structures and functional features. A copper-thioether axial bond shorter than the copper-thiolate bond is responsible of the electronic structure features, in contrast of all other natural orchineral sites where the copper thiolate bond is short. These sites display highly unusual features, such as: (1) a high reduction potential despite a strong interaction with the axial ligand, that weat tribute to changes in the hydrogen bond network, and (2) the ability to bind exogenous ligands such as imidazole and azide. This strategy widens the possibility of using natural protein scaffolds with functional features not present in nature. p { margin-bottom: 0.25cm; line-height: 120%; }