Solvent dependent and independent motions of CO–horseradish peroxidase examined by infrared spectroscopy and molecular dynamics calculations

• Published in: Biophysical chemistry Vol. 106; no. 1; pp. 1 - 14
• Main Authors: Kaposi, Andras D, Prabhu, Ninad V, Dalosto, Sergio D, Sharp, Kim A, Wright, W.W, Stavrov, Solomon S, Vanderkooi, J.M
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2003
• Subjects: Arginine - chemistry; Arginine - metabolism; Benzohydroxamic acid; Binding Sites; Carbon Monoxide - chemistry; Carbon Monoxide - metabolism; Carbonmonoxide; Heme - chemistry; Heme - metabolism; Histidine - chemistry; Histidine - metabolism; Horseradish peroxidase; Horseradish Peroxidase - chemistry; Horseradish Peroxidase - metabolism; Hydrogen-Ion Concentration; Hydroxamic Acids - chemistry; Hydroxamic Acids - pharmacology; Models, Molecular; Movement; Solvents - chemistry; Spectrophotometry, Infrared; Sugar glass; Temperature; Trehalose; Sugar glass; RMSD, root mean square deviation; Trehalose; Carbonmonoxide; BHA, benzohydroxamic acid; Benzohydroxamic acid; Horseradish peroxidase; HRP, horseradish peroxidase type III; IR, infrared; FWHM, full width at half maximum
• ISSN: 0301-4622; 1873-4200
• DOI: 10.1016/S0301-4622(03)00122-4

The role of the solvent matrix in affecting CO bound to ferrous horseradish peroxidase was examined by comparing band-widths of ν CO for the protein in aqueous solutions and in trehalose/sucrose glasses. We have previously observed that the optical absorption band and the CO stretching mode respond to the glass transition of glycerol/water in ways that depend upon the presence of substrate (Biochemistry 40 (2001) 3483). It is now demonstrated that the CO group band-width for the protein with bound inhibitor benzhydroxamic acid is relatively insensitive to temperature or the glass transition of the solvent. In contrast, in the absence of inhibitor, the band-width varies with the temperature that the glass is formed. The results show that solvent dependent and independent motions can be distinguished, and that the presence of substrate changes the protein such that the Fe–CO site is occluded from the solvent conditions. Molecular dynamic calculations, based upon X-ray structures, showed that the presence of benzhydroxamic acid decreases the distance between His42 and Arg38 and this leads for closer distances to the O of the CO from these residues. These results are invoked to account for the observed line width changes of the CO band.