Molecular cloning and nucleotide sequence of the α and β subunits of allophycocyanin from the cyanelle genome of Cyanophora paradoxa

Donald Ashley Bryant, R. de Lorimier, D. H. Lambert, J. M. Dubbs, V. L. Stirewalt, S. E. Stevens, Ronald D. Porter, J. Tam, E. Jay

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Abstract

The genes for the α- and β-subunit apoproteins of allophycocyanin (AP) were isolated from the cyanelle genome of Cyanophora paradoxa and subjected to nucleotide sequence analysis. The AP β-subunit apoprotein gene was localized to a 7.8-kilobase-pair Pst I restriction fragment from cyanelle DNA by hybridization with a tetradecameric oligonucleotide probe. Sequence analysis using that oligonucleotide and its complement as primers for the dideoxy chain-termination sequencing method confirmed the presence of both AP α- and β-subunit genes on this restriction fragment. Additional oligonucleotide primers were synthesized as sequencing progressed and were used to determine rapidly the nucleotide sequence of a 1336-base-pair region of this cloned fragment. This strategy allowed the sequencing to be completed without a detailed restriction map and without extensive and time-consuming subcloning. The sequenced region contains two open reading frames whose deduced amino acid sequences are 81-85% homologous to cyanobacterial and red algar AP subunits whose amino acid sequences have been determined. The two open reading frames are in the same orientation and are separated by 39 base pairs. AP α is 5' to AP β and both coding sequences are preceded by a polypurine, Shine-Dalgarno-type sequence. Sequences upstream from AP α closely resemble the Escherichia coli consensus promoter sequences and also show considerable homology to promoter sequences for several chloroplast-encoded psbA genes. A 56-base-pair palindromic sequence downstream from the AP β gene could play a role in the termination of transcription or translation. The allophycocyanin apoprotein subunit genes are located on the large single-copy region of the cyanelle genome.

Original languageEnglish (US)
Pages (from-to)3242-3246
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume82
Issue number10
DOIs
Publication statusPublished - Oct 2 1985

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