CpcM Posttranslationally Methylates Asparagine-71/72 of Phycobiliprotein Beta Subunits in Synechococcus sp. Strain PCC 7002 and Synechocystis sp. Strain PCC 6803

Abstract

Cyanobacteria produce phycobilisomes, which are macromolecular light-harvesting complexes mostly assembled from phycobiliproteins. Phycobiliprotein beta subunits contain a highly conserved ␥-N-methylasparagine residue, which results from the posttranslational modification of Asn71/72. Through comparative genomic analyses, we identified a gene, denoted cpcM, that (i) encodes a protein with sequence similarity to other S-adenosylmethionine-dependent methyltransferases, (ii) is found in all sequenced cyanobacterial genomes, and (iii) often occurs near genes encoding phycobiliproteins in cyanobacterial genomes. The cpcM genes of Synechococcus sp. strain PCC 7002 and Synechocystis sp. strain PCC 6803 were insertionally inactivated. Mass spectrometric analyses of phycobiliproteins isolated from the mutants confirmed that the CpcB, ApcB, and ApcF were 14 Da lighter than their wild-type counterparts. Trypsin digestion and mass analyses of phycobiliproteins isolated from the mutants showed that tryptic peptides from phycocyanin that included Asn72 were also 14 Da lighter than the equivalent peptides from wild-type strains. Thus, CpcM is the methyltransferase that modifies the amide nitrogen of Asn71/72 of CpcB, ApcB, and ApcF. When cells were grown at low light intensity, the cpcM mutants were phenotypically similar to the wild-type strains. However, the mutants were sensitive to high-light stress, and the cpcM mutant of Synechocystis sp. strain PCC 6803 was unable to grow at moderately high light intensities. Fluorescence emission measurements showed that the ability to perform state transitions was impaired in the cpcM mutants and suggested that energy transfer from phycobiliproteins to the photosystems was also less efficient. The possible functions of asparagine N methylation of phycobiliproteins are discussed.

Phycourobilin in Trichromatic Phycocyanin from Oceanic Cyanobacteria Is Formed Post-translationally by a Phycoerythrobilin Lyase-Isomerase

“…Only after maturation can phycobiliproteins form spontaneously trimers, which are then integrated into the PBSs by interaction with specific structural proteins, so-called linker polypeptides. These post-translational modifications reactions include the covalent attachment of 1-4 chromophores to each individual apoprotein (11)(12)(13)(14)(15)(16)(17)(18)(19)(20), methylation of an asparagine residue (21)(22)(23), and cleavage of N-terminal methionine residue (5, 24) (see UniProtKB/Swiss-Prot entry Q1XDQ2).…”

Biosynthesis of Cyanobacterial Phycobiliproteins in Escherichia coli : Chromophorylation Efficiency and Specificity of All Bilin Lyases from Synechococcus sp. Strain PCC 7002

“…This modification is thought to change the environment of the chromophore at position ␤-82 to minimize the rates of nonradiative energy loss within PBS (59,60). However, characterization of cpcM mutants also showed that these strains contain very high levels of reactive oxygen species (51).…”

“…While chromophore addition is the one type of posttranslational modification that all PBPs undergo, a second type of modification is a methylation reaction that occurs specifically on the ␤ subunits of most phycobiliproteins and is catalyzed by an S-adenosylmethionine-dependent methyltransferase designated CpcM (41,51,57). The methyltransferase reaction pro-duces a highly conserved ␥-N-methylasparagine residue at the ␤-72 position of almost all ␤-subunits isolated from cyanobacteria, red algae, and cryptomonads (17,35,36,45,65).…”

A preliminary analysis of the gene all0012 of Anabaena PCC 7120

“…During the maturation of PBP, another unique posttranslational modification occurs, the methylation of N-asparagine, which is located at the 72 position (consensus numbering), and no such modification has been found on the homologous position (Swanson and Glazer, 1990;Miller et al, 2008;Shen et al, 2008). Minami et al first reported that ApcB of Anabaena cylindria contained a modified aspartate residue at position 71 (Minami et al, 1985).…”

“…PCC 6803 methylated CpcB, ApcB, and ApcF. Even when the Asn72 of CpcB was replaced by Gln72, methylation occurred (Miller et al, 2008;Shen et al, 2008).…”