Low-temperature EPR and Mössbauer spectroscopy 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 g max below 3.2, and those featuring large g anisotropy with g max between 3.3 and 3.8, also denoted as highly axial low spin (HALS) species. Herein we present the detailed magnetic properties of cytochrome c 553 from Bacillus pasteurii (g max 3.36) and cytochrome c 552 from Nitrosomonas europaea (g max 3.34) over the pH range 6.2 to 8.2. Besides being structurally very similar, cytochrome C 553 shows the presence of a minor rhombic species at pH 6.2 (6%), whereas cytochrome c 552 has about 25% rhombic species over pH 7.5. The detailed MOssbauer analysis of cytochrome c 552 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 Mossbauer data of axial cytochromes c with His-Met iron coordination are consistent with an electronic (d xy ) 2 (d xz ) 2 (d yz ) 1 ground state, which is typical for Type I model hemes.