posted on 2015-06-02, 00:00authored byAmy E. Servid, Alison
L. McKay, Cherry A. Davis, Elizabeth M. Garton, Andreea Manole, Paul S. Dobbin, Michael
A. Hough, Colin R. Andrew
Five-coordinate
heme nitrosyl complexes (5cNO) underpin biological
heme-NO signal transduction. Bacterial cytochromes c′ are some of the few structurally characterized 5cNO proteins, exhibiting
a distal to proximal 5cNO transition of relevance to NO sensing. Establishing
how 5cNO coordination (distal vs proximal) depends on the heme environment
is important for understanding this process. Recent 5cNO crystal structures
of Alcaligenes xylosoxidans cytochrome c′ (AXCP) and Shewanella frigidimarina cytochrome c′ (SFCP) show a basic residue (Arg124 and Lys126,
respectively) near the proximal NO binding sites. Using resonance
Raman (RR) spectroscopy, we show that structurally characterized 5cNO
complexes of AXCP variants and SFCP exhibit a range of ν(NO)
(1651–1671 cm–1) and ν(FeNO) (519–536
cm–1) vibrational frequencies, depending on the
nature of the proximal heme pocket and the sample temperature. While
the AXCP Arg124 residue appears to have little impact on 5cNO vibrations,
the ν(NO) and ν(FeNO) frequencies of the R124K variant
are consistent with (electrostatically) enhanced Fe(II) → (NO)π*
backbonding. Notably, RR frequencies for SFCP and R124A AXCP are significantly
displaced from the backbonding trendline, which in light of recent
crystallographic data and density functional theory modeling may reflect
changes in the Fe–N–O angle and/or extent of σ-donation
from the NO(π*) to the Fe(II) (dz2) orbital. For R124A AXCP, correlation of vibrational
and crystallographic data is complicated by distal and proximal 5cNO
populations. Overall, this study highlights the complex structure–vibrational
relationships of 5cNO proteins that allow RR spectra to distinguish
5cNO coordination in certain electrostatic and steric environments.