The effects of some tumor markers on human erythrocyte (HCA-I and HCA-II), bovine erythrocyte (BCA) and bovine lung (CA-IV) carbonic anhydrase enzyme activities in vitro.

OBJECTIVE: The influence of prostatic acid phosphatase (PAP) and human chorionic gonadotropin (HCG), tumor markers have been investigated on human erythrocyte carbonic anhydrase (HCA-I and HCA-II) and bovine erythrocyte (BCA) and bovine lung carbonic anhydrase (CA-IV) in vitro. BACKGROUND: Tumor markers are substances that can often be detected in higher-than-normal amounts in the blood, urine, or body tissues of some patients with certain types of cancer. Tumor markers are produced either by the tumor itself or by the body in response to the presence of cancer or certain benign (noncancerous) conditions. In addition to their role in cancer diagnosis, some tumor marker levels are measured before treatment to help doctors plan appropriate therapy. RESULTS AND CONCLUSION: All of the tumor markers were determined to have inhibition effect, on human CA-I, CA-II, bovine erythrocyte CA (BCA) and bovine lung CA-IV isoenzymes. The effect of each tumor marker on CA was investigated by Wilbur-Andersen method modified by Rickly et al Inhibition effects of two different tumor markers on human CA-I, CA-II, bovine erythrocyte CA (BCA) and bovine lung CA-IV isoenzymes were determined by using the CO2-Hydratase method by plotting activity % vs (tumor markers). I50 values of tumor markers exhibiting inhibition effects were found by means of these graphs (Tab.1, Fig. 2, Ref. 20).

The activities of carbonic anhydrase and alkaline phosphatase in ancient human bones. Purification and characterization of outer peripheral, cytosolic, inner peripheral, and integral CA.

In the present study, bone carbonic anhydrase was isolated from ancient human bones and its characteristic features were determined. For this purpose, the skull bone of about 3000 years age was used. The purification was performed in four steps. Four different isoenzymes of CA, including outer peripheral, inner peripheral, integral, and cytosolic were purified and characterized. Affinity chromatography using Sepharose-4B-L-tyrosyn sulfanilamide as a support material was used in its purification. Two different methods were used for enzymatic activity determination: a) hydratase, and b) esterase methods. Bradford and Coomassie Brillant Blue methods were used for protein determination. Optimal pH, temperature, and molecular weight determinations were performed by conventional methods. The purification degree and the subunits, if present, were determined by SDS-PAGE. The effects of some chemicals on the enzyme were also investigated. The most cardinal finding was that the enzymatic activity has been found in antique human bone, showing some other enzymatic activity. That the alkaline phosphatase activity has been determined in the same sample supports the finding of carbonic anhydrase.

Carbonic anhydrase from bovine bone.

In this research, carbonic anhydrase enzyme, which was taken from the bones of an animal, was purified and characterized for the first time. For this, the bones of a young cow were used. The purification treatment was completed in three steps. Three different isoenzymes, such as peripheral, cystolic, and integral from the bone-cell cytozolic isoenzyme were purified and characterized. In purification of the three isoenzymes, the technique of affinity chromatography, which utilized Sepharose-4B-L-Tyrosine-Sulphanylamide, was used. In measuring the activities of enzymes, two different methods were applied. These are the esterase methods that utilize hydratase and p-nitrophenylacetate as substrate. The measurement of proteins was done with the methods of Bradford and Coomassie Brillant Blue. The optimum pH and temperature of each enzyme were measured and molecular weights were measured by gel-filtration. Its purity was examined by SDS-PAGE (3-10% alternating) electrophoresis and the inferior unit was defined. The inhibition effects of some chemicals were tested for each of the three isoenzymes.

Purification and characterization of carbonic anhydrase from bovine erythrocyte plasma membrane.

Carbonic anhydrase (CA) was purified from bovine erythrocyte plasma membrane and characterized in this study. For this purpose, the blood taken from young animals was hemolysed, the membrane fraction was separated, and this fraction was repeatedly washed. The enzyme (CA) was removed from the membrane with buffered TritonX-100 (1%); it could be purified at a factor of 22.8 by affinity chromatography. The CA obtained from erythrocyte membrane has an esterase activity as well as hydratase activity. The Vmax and Km of the enzyme for the substrate (p-nitrophenyl acetate) are 1.948x10(-3) mM/L x dak, and 3.596 mM, respectively. The purification degree of the enzyme was controlled by SDS-PAGE (3-10), which showed two distinct bands. It was determined that the enzyme had activity within the pH range of 4.5-9.5 and that the optimal pH was 7.5. The temperature at which it showed activity was 20-60 degrees C and optimal temperature was 37 degrees C. Molecular weight of CA was found to be 29844 and 61706 Dalton by gel filtration. On the other hand, sulfanilamide and acetazolamide affected the enzyme.