Membrane-associated carbonic anhydrase from rat lung. Purification, characterization, tissue distribution, and comparison with carbonic anhydrase IVs of other mammals.
Carbonic anhydrase (CA) IV was purified to homogeneity from rat lung microsomal and plasma membranes. The single N-terminal amino acid sequence showed 55% similarity to that reported for human CA IV. A monospecific antibody to the 39-kDa rat enzyme that cross-reacts on Western blots with CA IVs from other mammalian species was produced in rabbits. Digestion of rat lung enzyme with endoglycosidase (peptide-N-glycosidase F) reduced the Mr to 36,000, suggesting that rat CA contains one N-linked oligosaccharide chain. All of eight additional mammalian CA IVs that were examined also contained oligosaccharide chains, as evidenced by reduction in Mr from 52,000 (cow, sheep, and rabbit), 42,000 (pig, guinea pig, and dog), and 39,000 (mouse and hamster) to 36,000 after treatment of the respective lung microsomal membranes with peptide-N-glycosidase F. The 36-kDa human enzyme showed no change in molecular mass with this treatment. Thus, the human CA IV is the exceptional one in lacking carbohydrate. Rat lung CA IV was found to be relatively resistant to sodium dodecyl sulfate and to be anchored to membranes by a phosphatidylinositol-glycan linkage; both properties were found to be shared by other mammalian CA IVs. Western blot analysis indicated distribution of CA IV in rat tissues other than kidney and lung where it was previously known to be present. CA IV was particularly abundant in rat brain, muscle, heart, and liver, all locations where the CA IV enzyme was not known to be present previously. None was detected in rat skin or spleen.
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Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University School of Medicine, Missouri 63104.
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D000595:Amino Acid Sequence; D000818:Animals; D002256:Carbonic Anhydrases / Q000235:genetics / Q000302:isolation & purification*; D002462:Cell Membrane / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D008168:Lung / Q000201:enzymology*; D008322:Mammals; D008969:Molecular Sequence Data; D008970:Molecular Weight; D051381:Rats; D012689:Sequence Homology, Nucleic Acid
D007527:Isoenzymes; D002256:Carbonic Anhydrases