Interactions between hemoglobin and organic phosphates investigated with 31P nuclear magnetic resonance spectroscopy and ultrafiltration.
1. The chemical shifts (delta) of the phosphates of 2,3-diphosphoglycerate and adenosine triphosphate (ATP) were determined by phosphorus nuclear magnetic resonance (31P NMR) spectroscopy and were found to be displaced downfield following the addition of hemoglobin (3 mM) to a solution of either diphosphoglycerate (5 mM) or ATP (1 mM). 2. The binding of these compounds to hemoglobin was also determined by membrane ultrafiltration. A direct relationship was observed between the change in chemical shift ((delta delta) of the 2-P and 3-P of diphosphoglycerate and the percent diphosphoglycerate bound, when the latter was varied by altering pH, oxygenation state, or total diphosphoglycerate concentration. 3. In comparable studies with ATP binding, a linear relationship between the delta delta values of the gamma-, beta-, and alpha-P of ATP and the percent of ATP bound was not observed when the data from all of the experiments were plotted. NMR signals were not detectible in deoxyhemoglobin solutions containing 1 mM ATP but were seen in solutions containing 3.8 mM ATP. 4. The results indicate that 31P NMR spectroscopy is a promising tool for investigating organic phosphate interactions with hemoglobin.
D000255:Adenosine Triphosphate / Q000097:blood*; D001665:Binding Sites; D004163:Diphosphoglyceric Acids / Q000097:blood*; D006454:Hemoglobins; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010108:Oxyhemoglobins; D011485:Protein Binding; D011487:Protein Conformation; D014462:Ultrafiltration
D004163:Diphosphoglyceric Acids; D006454:Hemoglobins; D010108:Oxyhemoglobins; D000255:Adenosine Triphosphate