Molecular Pharmacology, Vol 16, 1065-1074, Copyright © 1979 by the American Society for Pharmacology and Experimental Therapeutics

Spectral Studies on the Interaction of Dichloromethane with Hemoglobin

¹ Departments of Pharmaceutical Chemistry and Pharmacology, University of California, San Francisco, California 94143

In spectrophotometric studies of hemoglobin solutions dichloromethane caused a decrease in the affinity and the cooperativity of oxygen binding. The maximum effects observed were at a dichloromethane partial pressure of 50 torr with no significant effects below 25 torr. At 50 torr dichloromethane oxygen affinity was decreased 50% and Hill n values, used as a measure of cooperativity, decreased from 2.95 to 2.07. Dichloromethane affected the binding of carbon monoxide to hemoglobin in solution as described by changes in Haldane " M" values, a measure of the relative affinities of carbon monoxide and oxygen. Carbon monoxide binding was decreased by dichloromethane partial pressures as low as 0.03 torr. Maximum effects, involving a change in M from 257 to 143, occurred at 5 torr dichloromethane, a concentration that did not affect oxygen binding to hemoglobin. Changes in the proton NMR spectrum for hemoglobin solutions in the presence of dichloromethane were compared to the results of the oxygen and carbon monoxide binding studies and to information on the binding sites for dichloromethane determined by X-ray crystallography. The contact shifted resonance attributed to the -heme was shifted approximately 1-2 ppm downfield in the presence of 100 torn dichloromethane. The upfield ring-current shifted resonance at +6.54 ppm, which has contributions from both the - and -E11 valine, was split into two peaks. Changes were also observed in the aromatic and methyl regions of the spectrum, suggestive of conformational changes in the protein with dichloromethane.

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ACKNOWLEDGMENT The authors wish to thank Dr. B. Schoenborn for his advice and encouragement early in the project, Dr. G. Matson for his help with NMR instrumentation and Dr. J. Trudell for his constructive criticism of the manuscript.