雑誌会（200115）回答_藤田

Axial interactions in the mixed-valent Cu_A active site and role of the axial methionine in electron transfer

1. http://www.pnas.org/content/110/36/14658.full.pdf#page=1&view=FitH

1. Ming-Li Tsai^a, 
2. Ryan G. Hadt^a, 
3. Nicholas M. Marshall^b, 
4. Tiffany D. Wilson^b, 
5. Yi Lu^b,¹, and
6. Edward I. Solomon^a,¹

Author Affiliations

1. Contributed by Edward I. Solomon, July 30, 2013 (sent for review May 31, 2013)

Significance

Long-range electron transfer (ET) is vital in energy transduction pathways. Within metalloprotein ET active sites, the role of the axial ligand in the mononuclear, blue copper (BC), also called type 1 Cu, sites is well defined, whereas its role in the binuclear mixed-valent Cu_A sites is less clear. This study defines the axial interaction in the mixed-valent binuclear Cu_A active site and its role in ET. The axial S(Met) ligand is essential in tuning down the reduction potential while not increasing the inner-sphere reorganization energy, a similar role to that found for the S(Met) ligand in BC. Furthermore, much like BC, the S(Met) bond in Cu_A is weak and therefore under entatic control by the surrounding protein matrix.

Abstract

Within Cu-containing electron transfer active sites, the role of the axial ligand in type 1 sites is well defined, yet its role in the binuclear mixed-valent Cu_A sites is less clear. Recently, the mutation of the axial Met to Leu in a Cu_A site engineered into azurin (Cu_A Az) was found to have a limited effect on E⁰ relative to this mutation in blue copper (BC). Detailed low-temperature absorption and magnetic circular dichroism, resonance Raman, and electron paramagnetic resonance studies on Cu_A Az (WT) and its M123X (X = Q, L, H) axial ligand variants indicated stronger axial ligation in M123L/H. Spectroscopically validated density functional theory calculations show that the smaller ΔE⁰ is attributed to H₂O coordination to the Cu center in the M123L mutant in Cu_A but not in the equivalent BC variant. The comparable stabilization energy of the oxidized over the reduced state in Cu_A and BC (Cu_A ∼ 180 mV; BC ∼ 250 mV) indicates that the S(Met) influences E⁰ similarly in both. Electron delocalization over two Cu centers in Cu_A was found to minimize the Jahn–Teller distortion induced by the axial Met ligand and lower the inner-sphere reorganization energy. The Cu–S(Met) bond in oxidized Cu_A is weak (5.2 kcal/mol) but energetically similar to that of BC, which demonstrates that the protein matrix also serves an entatic role in keeping the Met bound to the active site to tune down E⁰ while maintaining a low reorganization energy required for rapid electron transfer under physiological conditions.