INFRARED ABSORPTION STUDY OF THE HEME POCKET DYNAMICS OF
CARBONMONOXYHEME PROTEINS
Andras D Kaposi 1, Jane M Vanderkooi 2 and Solomon S. Stavrov 3*
1 Institute of Biophysics and Radiation Biology, Semmelweis University of Medicine
2 Univ. of Pennsylvania School of Medicine
3 Tel Aviv Univ; Sackler School of Medicine
* To whom correspondence should be addressed. E-mail: stavrov{at}post.tau.ac.il.
Submitted on June 8, 2005
Revised on July 13, 2005
Accepted on 23 August 2006
 |
Abstract |
|---|
The temperature dependences of the infrared absorption CO bands of carboxy complexes of horseradish peroxidase (HRP(CO)) in glycerol/water mixture at pH 6.0 and 9.3 are interpreted using the theory of optical absorption bandshape. The bands' anharmonic behavior is explained assuming that there is higher-energy set of conformational substates (CSSh), which are populated upon heating and correspond to the protein substates with disordered water molecule in the heme pocket. Analysis of the second moments of the CO bands of the carboxy complexes of myoglobin (Mb(CO)) and hemoglobin (Hb(CO)), and of HRP(CO) with benzohydroxamic acid (HRP(CO)+BHA) shows that the low energy CSSh exists also in the "open" conformation of Mb(CO), where the heme pocket is spacious enough to accommodate a water molecule. In the HRP(CO)+BHA and "closed" conformations of Mb(CO) and Hb(CO) the heme pocket is packed with BHA and different amino acids, the CSSh has much higher energy and is hardly populated even at the highest temperatures. Therefore only motions of these amino acids contribute to the band broadening. These motions are linked to the protein surface and frozen in the glassy matrix, whereas in the liquid solvent they are harmonic. Thus the second moment of the CO band is temperature independent in glass and is proportional to the temperature in liquid. The temperature dependence of the second moment of the CO peak of HRP(CO) in the trehalose glass exhibits linear coupling to an oscillator. This oscillator can be a moving water molecule locked in the heme pocket in the whole interval of temperatures or a trehalose molecule located in the heme pocket.
Key Words:
conformational substate, glass-liquid transition, hemoglobin, horseradish peroxidase, myoglobin, protein dynamics
