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.
Demir
Y
Y
Atatürk Universitesi, Fen-Edebiyat Fakültesi, Erzurum, Turkey. demirn@yahoo.com
Demir
N
N
Yildirim
S
S
Nadaroglu
H
H
Karaosmanoglu
M
M
Bakan
E
E
D000469:Alkaline Phosphatase / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D001842:Bone and Bones / Q000166:cytology / Q000201:enzymology*; D002256:Carbonic Anhydrases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D002417:Cattle; D003600:Cytosol / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D005580:Fossils; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D013696:Temperature; D014421:Turkey
D007527:Isoenzymes; D000469:Alkaline Phosphatase; D002256:Carbonic Anhydrases