Human mitochondrial carbonic anhydrase: cDNA cloning, expression, subcellular localization, and mapping to chromosome 16.
A full-length cDNA clone encoding human mitochondrial carbonic anhydrase (CA), CA V, was isolated from a human liver cDNA library. The 1123-bp cDNA includes a 55-bp 5' untranslated region, a 915-bp open reading frame, and a 153-bp 3' untranslated region. Expression of the cDNA in COS cells produced active enzyme. The 34-kDa precursor and 30-kDa mature form of CA V were identified on Western blots of COS-cell homogenates by a CA V-specific antibody raised to a synthetic peptide corresponding to the C-terminal 17 aa of CA V. Both 34-kDa and 30-kDa bands were also present in mitochondria isolated from transfected COS cells, whereas only the 30-kDa band was present in mitochondria isolated from normal human liver. The N-terminal sequence determined directly on the 30-kDa soluble CA purified from transfected COS cells indicated that processing of the precursor to mature human CA V involves removal of a 38-aa mitochondrial leader sequence. The 267-aa sequence deduced for mature human CA V shows 30-49% similarity to amino acid sequences of previously characterized human CAs (CA I-CA VII) and 76% similarity to the corresponding amino acid sequence deduced from the mouse cDNA. PCR analysis of DNAs from human-rodent somatic cell hybrids localized the gene for CA V to human chromosome 16, the same chromosome to which CA VII has previously been mapped.
Nagao
Y
Y
Edward A. Doisy Department of Biochemistry and Molecular Biology, St. Louis University School of Medicine, MO 63104.
Platero
J S
JS
Waheed
A
A
Sly
W S
WS
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002256:Carbonic Anhydrases / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002874:Chromosome Mapping; D002885:Chromosomes, Human, Pair 16; D003001:Cloning, Molecular / Q000379:methods; D004247:DNA / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D008930:Mitochondria, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction / Q000379:methods; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D004247:DNA; D002256:Carbonic Anhydrases