Human mitochondrial carbonic anhydrase: cDNA cloning, expression, subcellular localization, and mapping to chromosome 16

Abstract

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.