α and β Subunits of Cyanidium caldarium phycocyanin. Properties and amino acid sequences at the amino terminus

Abstract

Phycocyanin was isolated and purified from the unicellular alga, Cyanidium caldarium. Subunits were prepared on a Bio-Rex-70 column developed stepwise with urea solutions (pH 1.9). The alpha subunit eluted in 8 M urea and the beta subunit eluted in 9 M urea. The alpha and beta subunits displayed absorption maxima at 660, 354, and 277 nm in 8 M and 9 M urea. The alpha:beta ratio of total absorbance under the 660-nm peak was 0.56 suggesting an alpha:beta phycocyanobilin chromophore ration of 1:2. On calibrated sodium dodecyl sulfate gels, the alphs subunit had an estimated molecular weight of 15,500 plus or minus 1100 and the beta subunit has an estimated molecular weight of 18,300 plus or minus 300. Minimum molecular weights based on one histidine residue per subunit were 16,300 for the alpha subunit and 18,750 for the beta subunit. Phycocyanin displayed a single visible absorption maximum at 625 nm and two positive circular dichroic bands at 632 and 610 nm. The alpha and beta subunits displayed single visible absorption maxima at 618 and 600 nm and single positive circular dichroic peaks at 620 and 585 nm, respectively. Two-dimensional maps of tryptic digests of the alpha and beta subunits revealed distinct patterns of peptides each of which was consistent with the lysine and arginine composition of these polypeptides. Maps of tryptic digests of phycocyanin contained 25 major peptides (a total of 27 lysine and arginine residues). Automated sequence analysis of separated subunits revealed a 70% homology within the first 27 residues at the amino terminus of the alpha and beta subunits of C. caldarium phycocyanin.

Genes encoding major light-harvesting polypeptides are clustered on the genome of the cyanobacterium Fremyella diplosiphon.

“…Each phycobiliprotein is composed of two related subunits, a and ,B, to which one or more tetrapyrole chromophores are attached (4), conferring upon each phycobiliprotein a characteristic light-absorption and emission spectrum. Amino acid (6)(7)(8)(9)(10)(11)(12)(13) and DNA (14)(15)(16) sequence data show considerable sequence conservation among the various phycobiliproteins (APC, PC, and phycoerythrin) as well as between the a and P3 subunits of a given phycobiliprotein. These findings suggest that phycobiliprotein genes arose via duplications of an ancestral sequence (8).…”

Bilin—apoprotein linkages in rhodophytan phycobiliproteins: The role of cysteine

“…Cyanidium caldarium cells were grown in medium containing inorganic salts as described previously [9] except that 10 mCi of HZ3'S04 (New Engl. Nuclear, Boston, MA) was added to the medium and HzS04 was omitted.…”

“…Nuclear, Boston, MA) was added to the medium and HzS04 was omitted. Radiolabeled phycocyanin and allophycocyanin were isolated from algal cells and purified as described [9,10] . The a and /3 subunits of phycocyanin were prepared by ion-exchange chromatography on Bio-Rex 70 columns according to the method of Glazer buffer was added to make the final iodoacetamide concentration 0.1 M. The reaction mixtures were incubated for an additional 30 min at room temperature under nitrogen, excess mercaptoethanol was added to terminate the reaction, and the alkylated phycobiliproteins were dialyzed against distilled water and lyophilized.…”

“…We have shown that phycocyanin and allophycocyanin from the unicellular rhodophyte, Cyanidium caldarium, are comprised of dissimilar polypeptide subunits, termed (Y and 0, that the OL : fl chromophore composition of the phycocyanin subunits is 1 : 2, and that the Q : @ chromophore composition of the allophycocyanin subunits is 1 : 1 [9,10] . Clearly the complexity of the subunit and chromophore composition of these phycobiliproteins could allow for more than one type of covalent attachment between phycocyanobilin and amino acids in the subunit polypeptides.…”

Isolation and characterization of a gene for a major light-harvesting polypeptide from Cyanophora paradoxa

“…Each of the phycobiliproteins contains an a and P3 subunit and each subunit covalently binds one or more tetrapyrrole chromophores (1). Amino acid sequence data (9)(10)(11)(12)(13)(14)(15)(16)(17) have shown amino acid conservation within the a subunits as a group and the f3 subunits as a group and among the various a and (3 subunits. This had led to speculation that individual polypeptide subunits of the different phycobiliprotein groups may have evolved from duplications of a single progenitor gene (11).…”