Low temperature EPR and Mossbauer spectroscopic of two cytochromes with His-Met axial coordination exhibiting HALS signals

C-type cytochromes with histidine–methionine (His–Met) iron coordination play important roles in electron-transfer reactions and in enzymes. Low-temperature electron paramagnetic resonance (EPR) spectra of low-spin ferric cytochromes c can be divided into two groups, depending on the spread of g values: the normal rhombic ones with small g anisotropy and gmax below 3.2, and those featuring large g anisotropy with gmax between 3.3 and 3.8, also denoted as highly axial low spin (HALS) species. Herein we present the detailed magnetic properties of cytochrome c553 from Bacillus pasteurii (gmax 3.36) and cytochrome c552 from Nitrosomonas europaea (gmax 3.34) over the pH range 6.2 to 8.2. Besides being structurally very similar, cytochrome c553 shows the presence of a minor rhombic species at pH 6.2 (6 %), whereas cytochrome c552 has about 25% rhombic species over pH 7.5. The detailed Moessbauer analysis of cytochrome c552 confirms the presence of these two low-spin ferric species (HALS and rhombic) together with an 8% ferrous form with parameters comparable to the horse cytochrome c. Both EPR and Moessbauer data of axial cytochromes c with His–Met iron coordination are consistent with an electronic (dxy)2 (dxz)2 (dyz)1 ground state, which is typical for Type I model hemes.