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cDNA cloning of the human retinal rod Na-Ca + K exchanger: comparison with a revised bovine sequence
J E Tucker1, R J Winkfein, C B Cooper
1Department of Medical Biochemistry, Faculty of Medicine, University of Calgary, Canada.
Purpose:
To clone the complementary DNA of the human retinal rod Na-Ca + K exchanger.
Methods:
A human retinal cDNA library was screened initially with a radiolabeled probe representing the entire bovine rod Na-Ca + K exchanger cDNA and subsequently with probes from polymerase chain reaction fragments of the human retinal rod Na-Ca + K exchanger obtained after the initial screen. Twelve positive clones were used to obtain the entire coding sequence of the human retinal rod Na-Ca + K exchanger.
Results:
The cDNA of the human retinal rod Na-Ca + K exchanger codes for a protein of 1081 amino acids, which shows 64.3% overall identity with the bovine retinal rod Na-Ca + K exchanger at the amino acid level. The two sets of putative transmembrane-spanning domains and their short connecting loops showed the highest degree of identity (94%-95%), whereas the extracellular loop at the N terminus showed a 59% identity. The large cytosolic loop that bisects the two sets of transmembrane-spanning domains contained two large deletions in the human exchanger; the first deletion contains 18 amino acids, whereas the second deletion involves a series of repeats that are dominated by acidic amino acid residues observed in the bovine, but not in the human, sequence. The authors observed that the bovine sequence contains a ninth repeat in addition to the eight repeats of the published sequence.
Conclusions:
The authors cloned the cDNA of the human retinal rod Na-Ca + K exchanger as a first step in examining the possibility that this gene could be the locus of disease-causing mutations.