Nitrite reductase activity within an antiparallel de novo scaffold

Catalysis and Electron Transfer in De Novo Designed Metalloproteins

“…Focus on long-distance, secondary sphere, and even primary sphere (ε vs δ nitrogen of His) effects have improved on the original rate up to 730-fold, but there is still a 3–4 order of magnitude rate gap compared to native CuNiR. ,, Most recently, a CuHis 3 site with several possible secondary sphere designs was incorporated into the asymmetric scaffold GRα 3 D, which showed that the CuNiR activity is sensitive to the metal site design and position within the scaffold (Figure ). Superoxide dismutase (SOD) and monooxygenase mimics have been worked on by several laboratories, and the Cu-only SOD constructs show surprising insensitivity to the usually observed correlation of redox potential and catalytic efficiency. , …”

“…245,250,254 Most recently, a CuHis 3 site with several possible secondary sphere designs was incorporated into the asymmetric scaffold GRα 3 D, which showed that the CuNiR activity is sensitive to the metal site design and position within the scaffold (Figure 34). 258 Superoxide dismutase (SOD) and monooxygenase mimics have been worked on by several laboratories, and the Cu-only SOD constructs show surprising insensitivity to the usually observed correlation of redox potential and catalytic efficiency. 259,290 The de novo design of monometal metalloenzymes is a strong showcase for the lessons we can learn about the simplest functional unit necessary for metalloenzyme activity.…”

“…Models were made using the crystal structural of GRα 3 D (PDB 6DS9). Reprinted with permission from ref . Copyright 2021 Springer Nature.…”

“…176 As was employed for carbonic anhydrase mimics (see section 3.1), the Pecoraro lab attempted to generate asymmetric environments around copper by incorporating a CuHis 3 site within the α 3 D and GRα 3 D scaffolds to investigate the effect of single Asp or His residues near the active site and the overall effect of large scaffold changes on CuNiR catalysis. 258 The same CuHis 3 site was 6 times more active when incorporated into α 3 D and up to 18 times more active in the more stable scaffold GRα 3 D. That the surrounding protein matrix could impact a metal center's function is not novel, but it had only recently been recognized in de novo metalloprotein design (see section 2.2). 61 The CuNiR activities of scaffolds containing Asp residues positioned above or below the CuHis 3 binding site or a fourth His residue were determined using a series of constructs previously investigated for CuSOD activity (see section 3.3).…”

A De Novo‐Designed Type 3 Copper Protein Tunes Catechol Substrate Recognition and Reactivity

“…This study represents a breakthrough in de novo metalloprotein design. Differently from T1 and T2 mononuclear copper sites, which have been widely constructed in a variety of de novo scaffolds, [4,44,[56][57][58][59][60][61] the T3 di-nuclear sites have been mainly studied in natural proteins [62] and in small inorganic complexes. [55,63] Very recently, Song and co-workers installed a di-copper site at the C 2 -symmetric interface of the hexameric acetyltransferase, using the unnatural bipyridyl-alanine as a tight chelating residue.…”