Shen Gaozhong, Leonard Heidi S, Schluchter Wendy M, Bryant Donald A
Department of Biochemistry and Molecular Biology, S-235 Frear Building, The Pennsylvania State University, University Park, PA 16802, USA.
J Bacteriol. 2008 Jul;190(14):4808-17. doi: 10.1128/JB.00436-08. Epub 2008 May 9.
Cyanobacteria produce phycobilisomes, which are macromolecular light-harvesting complexes mostly assembled from phycobiliproteins. Phycobiliprotein beta subunits contain a highly conserved gamma-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.
蓝细菌产生藻胆体,藻胆体是主要由藻胆蛋白组装而成的大分子捕光复合体。藻胆蛋白β亚基含有一个高度保守的γ-N-甲基天冬酰胺残基,该残基是天冬酰胺71/72的翻译后修饰产物。通过比较基因组分析,我们鉴定出一个名为cpcM的基因,该基因(i)编码一种与其他依赖S-腺苷甲硫氨酸的甲基转移酶具有序列相似性的蛋白质,(ii)存在于所有已测序的蓝细菌基因组中,并且(iii)在蓝细菌基因组中经常出现在编码藻胆蛋白的基因附近。聚球藻属菌株PCC 7002和集胞藻属菌株PCC 6803的cpcM基因被插入失活。对从突变体中分离的藻胆蛋白进行质谱分析证实,CpcB、ApcB和ApcF比其野生型对应物轻14 Da。对从突变体中分离的藻胆蛋白进行胰蛋白酶消化和质谱分析表明,来自包括天冬酰胺72的藻蓝蛋白的胰蛋白酶肽段也比来自野生型菌株的等效肽段轻14 Da。因此,CpcM是修饰CpcB、ApcB和ApcF的天冬酰胺71/72酰胺氮的甲基转移酶。当细胞在低光照强度下生长时,cpcM突变体在表型上与野生型菌株相似。然而,这些突变体对高光胁迫敏感,集胞藻属菌株PCC 6803的cpcM突变体在中等高光强度下无法生长。荧光发射测量表明,cpcM突变体中进行状态转换的能力受损,这表明从藻胆蛋白到光系统的能量转移效率也较低。本文讨论了藻胆蛋白天冬酰胺N甲基化的可能功能。