Donnas K. Starkweather

The binding of sulfaphenazole to fetal, neonatal, and adult human plasma albumin

In man, sulfaphenazole has a lower affinity for the albumin of the fetus and neonate than for that of the adult. Removal of bound endogenous anions from neonatal albumin by treatment with charcoal at pH 3.0 restored the affinity of the protein for sulfaphenazole to the adult level. Prior treatment of adult albumin with bilirubin reduced the affinity of the protein for sulfaphenazole to that observed in fetal and neonatal albumins. Thus the reduced affinity of su1faphenazole for fetal and neonatal albumin compared to that for adult albumin probably is the result of the presence of a tightly bound endogenous ligand, such as bilirubin, on fetal and neonatal albumin.

The interaction of some spin-labeled sulfonamides with bovine erythrocyte carbonic anhydrase B☆

Abstract 1. 1. Spin-labeled analogs of acetazolamide and sulfanilamide containing the nitroxide free radical have been synthesized and their interaction with bovine erythrocyte carbonic anhydrase B (EC 4.2.1.1) studied by means of electron spin resonance. 2. 2. When 2,2,5,5-tetramethyl-3-[(5-sulfamoyl-1,3,4-thiadiazol-2-yl)carbamoyl]-1-pyrrolidinyloxyl bound to carbonic anhydrase, the nitroxide group of this spin label became highly immobilized at the enzyme active site. 3. 3. When other spin-labeled sulfonamides bound to bovine erythrocyte carbonic anhydrase B, it was found that the molecular motion of the nitroxide group was dependent upon the distance of this free radical from the aromatic sulfonamide group. 4. 4. When 2,2,5,5-tetramethyl-3-[( p -sulfamoyl)carbamoyl]propionamido]-1-pyrrolidinyloxyl bound to carbonic anhydrase, the nitroxide group no longer interacted with the active site. This suggests that the active site of bovine erythrocyte carbonic anhydrase B is a narrow cleft approx. 14.5 A in depth.

Species Differences in the Binding of Phenylbutazone to Plasma Albumin

The binding of phenylbutazone to plasma albumins from man, rat, rabbit and dog has been measured at 37°C by equilibrium dialysis. Species differences were found both in the number of phenylbutazone bi

A Novel Assay for Erythrocyte Carbonic Anhydrase and Certain Sulfonamide Drugs Employing Spin-Labels

A novel assay procedure has been developed for the determination of sulfonamide drugs with the general formula ArSO2NH2 where Ar = aryl (either homocyclic or heterocyclic). The assay depends on the displacement of a spin-labeled sulfonamide from the active site of bovine carbonic anhydrase B and can detect concentrations of sulfonamide as low as 0.3 µg/ml. This technique can also be used to assay low concentrations of carbonic anhydrase in hemolysates of red blood cells without prior removal of the hemoglobin or isolation of the enzyme.

A spin label study of human erythrocyte ghost membranes damaged by methyl phenyldiazenecarboxylate (azoester)

Abstract The mechanism of methyl phenyldiazenecarboxylate (azoester) damage to human erythrocyte membranes has been investigated by means of spin labels. Azoester treatment exposed protein binding sites for non-covalently bound stearic acid and androstane spin labels that were occult in the untreated membrane. Experiments with iodoacetamide and N-alkylmaleimide spin labels suggested that azoester destroyed membrane sulfhydryl groups. No change in the structural integrity of membrane lipid components could be detected.

A spin-label study of horse erythrocyte carbonic anhydrases C and D

Abstract 1. 1. The topography of the active sites of horse erythrocyte carbonic anhydrases C and D has been studied by means of a series of spin-labeled sulfanilamide analogs. 2. 2. The active site of horse carbonic anhydrase D is a narrow cleft approximately 14 A in depth. The active site of horse carbonic anhydrase C is similar to that of the D isozyme but somewhat deeper. 3. 3. A comparison of horse erythrocyte carbonic anhydrases with the human erythrocyte carbonic anhydrases B and C indicates that the low activity enzymes (horse D and human B) have narrower active sites than the higher activity isozymes (horse C and human C).