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dc.contributor.advisor Olson, John S.
dc.creatorLemon, Douglas Dale
dc.date.accessioned 2009-06-04T00:12:33Z
dc.date.available 2009-06-04T00:12:33Z
dc.date.issued 1989
dc.identifier.urihttps://hdl.handle.net/1911/16255
dc.description.abstract O$\sb2$ transport was examined by measuring the fractional saturation of concentrated hemoglobin solutions flowing through an artificial capillary (diameter $\approx$ 27 $\mu$m). The measured effects of pH, hemoglobin concentration, O$\sb2$ tension, temperature, and organic phosphate were analyzed by a mathematical model which included the geometry of the capillary, parabolic flow inside the lumen, and cooperative O$\sb2$ binding. Oxygen exchange was limited by diffusion and therefore governed by the magnitude of the O$\sb2$ gradient between the intracapillary fluid phase and the external gas space. In uptake experiments, O$\sb2$ flux was determined primarily by the external O$\sb2$ tension (160 mm Hg) because the internal O$\sb2$ pressure was kept small due to chemical combination with hemoglobin. In release experiments, the external O$\sb2$ tension was maintained at zero, and the transport rate was determined by the intracapillary oxygen partial pressure, which was proportional to the O$\sb2$ half-saturation pressure of the hemoglobin sample. Thus, factors that change the affinity of hemoglobin for oxygen, such as pH, temperature, and organic phosphate concentration, influence strongly the rate of O$\sb2$ release but have little effect on the rate of O$\sb2$ uptake. The rate-limiting step for erythrocyte CO$\sb2$ transport is the transmembrane exchange of Cl$\sp-$ and HCO$\sb3\sp-$ anions. This process was measured by following extracellular pH changes using a pH-sensitive fluorescent dye in a stopped-flow mixing device equipped with front-face optics. Initial pH and chloride gradients induced pH relaxations which were sensitive to the specific inhibitor 4,4$\prime$-diisothiocyano-2,2$\prime$-stilbenedisulfonic acid (DIDS). Special mixing experiments, designed to minimize anion competition for the transporter binding site(s), were simulated mathematically by models written for ping-pong and random ternary complex mechanisms. The experimentally observed rate dependence on initial extracellular bicarbonate concentration was approximated better by the ping-pong model, and the theoretically derived values for Cl$\sp-$ and HCO$\sb3\sp-$ binding (both K$\sb {\rm D}$'s = 5 mM) as well as the translocation rate (1.1 $\times$ 10$\sp4$ s$\sp{-1}$) agreed very well with literature values for inhibition constants and protein turnover number, respectively. Finally the simultaneous measurement of O$\sb2$ uptake and subsequent HCO$\sb3\sp-$/Cl$\sp-$ exchange was demonstrated. The hemoglobin color change upon oxygen binding caused a rapid change in fluorescence, followed by a slower, DIDS-sensitive pH equilibration.
dc.format.extent 198 p.
dc.format.mimetype application/pdf
dc.language.iso eng
dc.subjectBiochemistry
Physiology
dc.title Oxygen and carbon dioxide exchange by human hemoglobin and erythrocytes
dc.type.genre Thesis
dc.type.material Text
thesis.degree.department Chemistry
thesis.degree.discipline Natural Sciences
thesis.degree.grantor Rice University
thesis.degree.level Doctoral
thesis.degree.name Doctor of Philosophy
dc.identifier.citation Lemon, Douglas Dale. "Oxygen and carbon dioxide exchange by human hemoglobin and erythrocytes." (1989) Diss., Rice University. https://hdl.handle.net/1911/16255.


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