title: Comparative analysis of cryoprotective agents influence on thermodynamic and kinetic parameters of equine and human hemoglobin molecules.
abstract: BACKGROUND
  Critical to the understanding the mechanism of destruction and protection during cryopreservation of biological objects is the knowledge of the conformational transitions of biopolymers experiencing low temperatures in the presence of cryoprotective agents. This information may be derived from the kinetic and thermodynamic parameters of macromolecular thermal denaturation kinetics under different environmental conditions.
  
  
  OBJECTIVE
  The study deals with the influence of cryoprotective agents (glycerol, 1.2-propanediol (1.2-PD), and dimethyl sulfoxide (DMSO)) on thermodynamic and kinetic parameters of equine and human hemoglobin.
  
  
  METHODS
  Thermograms were recorded with differential scanning adiabatic microcalorimeter (DASM-4, Biopribor, Russia).
  
  
  RESULTS
  Temperatures and enthalpy changes in the denaturation of hemoglobins (ΔHcal) in the presence of cryoprotective agents from 0-40% (w/w) were determined. Within the whole concentration range glycerol slightly alters the hemoglobin denaturation temperature while DMSO and 1.2-PD lead to a significant decrease. The addition of cryoprotectants to hemoglobin solutions induces a complex behavior of ΔHcal concentration dependences of denaturation caused by intra- and intermolecular processes such as hydration of the porphyrin cycle, heme cleavage from heme-containing proteins, disorder in hydrophobic contacts with globin etc. These factors may decrease thermal stability by loosening hemoglobin molecules and spatial disruption of fragments of the protein. Activation energy of irreversible unfolding of equine hemoglobin at heating was estimated using the approach of Sanchez-Ruiz et al. DMSO and 1.2-PD decreased activation energy values.
  
  
  CONCLUSIONS
  The data indicate the reduction of protein thermal stability by DMSO and 1.2-PD. Glycerol slightly increases hemoglobin thermal stability within the studied range of concentrations.
authors.affiliation: Department of Cryobiophysics, Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Kharkov, Ukraine. alexazin@mail.ru and ju-st7@yandex.ua.
authors.forename: Aleksandra V
authors.initials: AV
authors.lastname: Zinchenko
authors.affiliation: Department of Cryobiophysics, Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Kharkov, Ukraine.
authors.forename: Yuliana S
authors.initials: YS
authors.lastname: Govorova
mesh_terms: D000818:Animals; D015925:Cryopreservation; D003451:Cryoprotective Agents / Q000737:chemistry*; D004121:Dimethyl Sulfoxide / Q000737:chemistry*; D005990:Glycerol / Q000737:chemistry*; D006454:Hemoglobins / Q000737:chemistry*; D006736:Horses; D006801:Humans; D007700:Kinetics; D019946:Propylene Glycol / Q000737:chemistry*; D011489:Protein Denaturation; D055550:Protein Stability; D013696:Temperature; D013816:Thermodynamics
chemical_list: D003451:Cryoprotective Agents; D006454:Hemoglobins; D019946:Propylene Glycol; D005990:Glycerol; D004121:Dimethyl Sulfoxide