Characterization of CA XV, a new GPI-anchored form of carbonic anhydrase.
The main function of CAs (carbonic anhydrases) is to participate in the regulation of acid-base balance. Although 12 active isoenzymes of this family had already been described, analyses of genomic databases suggested that there still exists another isoenzyme, CA XV. Sequence analyses were performed to identify those species that are likely to have an active form of this enzyme. Eight species had genomic sequences encoding CA XV, in which all the amino acid residues critical for CA activity are present. However, based on the sequence data, it was apparent that CA XV has become a non-processed pseudogene in humans and chimpanzees. RT-PCR (reverse transcriptase PCR) confirmed that humans do not express CA XV. In contrast, RT-PCR and in situ hybridization performed in mice showed positive expression in the kidney, brain and testis. A prediction of the mouse CA XV structure was performed. Phylogenetic analysis showed that mouse CA XV is related to CA IV. Therefore both of these enzymes were expressed in COS-7 cells and studied in parallel experiments. The results showed that CA XV shares several properties with CA IV, i.e. it is a glycosylated glycosylphosphatidylinositol-anchored membrane protein, and it binds CA inhibitor. The catalytic activity of CA XV is low, and the correct formation of disulphide bridges is important for the activity. Both specific and non-specific chaperones increase the production of active enzyme. The results suggest that CA XV is the first member of the alpha-CA gene family that is expressed in several species, but not in humans and chimpanzees.
Hilvo
Mika
M
Institute of Medical Technology, University of Tampere and Tampere University Hospital, Biokatu 6, 33520 Tampere, Finland. Mika.Hilvo@uta.fi
Tolvanen
Martti
M
Clark
Amy
A
Shen
Bairong
B
Shah
Gul N
GN
Waheed
Abdul
A
Halmi
Piia
P
Hänninen
Milla
M
Hämäläinen
Jonna M
JM
Vihinen
Mauno
M
Sly
William S
WS
Parkkila
Seppo
S
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002256:Carbonic Anhydrases / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002522:Chlorocebus aethiops; D019295:Computational Biology; D004220:Disulfides / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006031:Glycosylation; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D002679:Pan troglodytes / Q000235:genetics; D010802:Phylogeny; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D011544:Pseudogenes / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
D004220:Disulfides; D017261:Glycosylphosphatidylinositols; D011994:Recombinant Proteins; D002256:Carbonic Anhydrases; C507974:carbonic anhydrase XV, mouse