Department of Molecular Biology and Biochemistry, Rutgers University, Busch Campus, 604 Allison Road, Piscataway, NJ 08854-8082, USA.
Photosynth Res. 2011 Sep;108(2-3):201-14. doi: 10.1007/s11120-011-9681-x. Epub 2011 Aug 24.
In order to obtain an improved understanding of the assembly of the bacterial photosynthetic apparatus, we have conducted a proteomic analysis of pigment-protein complexes isolated from the purple bacterium Rhodobacter sphaeroides undergoing acclimation to reduced incident light intensity. Photoheterotrophically growing cells were shifted from 1,100 to 100 W/m(2) and intracytoplasmic membrane (ICM) vesicles isolated over 24-h were subjected to clear native polyacrylamide gel electrophoresis. Bands containing the LH2 and reaction center (RC)-LH1 complexes were excised and subjected to in-gel trypsin digestion followed by liquid chromatography (LC)-mass spectroscopy (MS)/MS. The results revealed that the LH2 band contained distinct levels of the LH2-α and -β polypeptides encoded by the two puc operons. Polypeptide subunits encoded by the puc2AB operon predominated under high light and in the early stages of acclimation to low light, while after 24 h, the puc1BAC components were most abundant. Surprisingly, the Puc2A polypeptide containing a 251 residue C-terminal extension not present in Puc1A, was a protein of major abundance. A predominance of Puc2A components in the LH2 complex formed at high light intensity is followed by a >2.5-fold enrichment in Puc1B levels between 3 and 24 h of acclimation, accompanied by a nearly twofold decrease in Puc2A levels. This indicates that the puc1BAC operon is under more stringent light control, thought to reflect differences in the puc1 upstream regulatory region. In contrast, elevated levels of Puc2 polypeptides were seen 48 h after the gratuitous induction of ICM formation at low aeration in the dark, while after 24 h of acclimation to low light, an absence of alterations in Puc polypeptide distributions was observed in the upper LH2-enriched gel band, despite an approximate twofold increase in overall LH2 levels. This is consistent with the origin of this band from a pool of LH2 laid down early in development that is distinct from subsequently assembled LH2-only domains, forming the LH2 gel band.
为了更好地了解细菌光合装置的组装过程，我们对适应低光照强度的球形红杆菌细胞内膜（ICM）囊泡中的色素蛋白复合物进行了蛋白质组分析。将异养生长的细胞从 1,100 瓦/平方米转移到 100 瓦/平方米，然后在 24 小时内分离 ICM 囊泡，并进行清晰的天然聚丙烯酰胺凝胶电泳。分离出含有 LH2 和反应中心（RC）-LH1 复合物的条带，并进行胶内胰蛋白酶消化，然后进行液相色谱（LC）-质谱（MS）/MS。结果表明，LH2 带含有由两个 puc 操纵子编码的 LH2-α和-β多肽的不同水平。在高光和低光适应的早期阶段，puc2AB 操纵子编码的多肽亚基占主导地位，而 24 小时后，puc1BAC 成分最为丰富。令人惊讶的是，Puc2A 多肽含有一个 251 个残基的 C 端延伸，不存在于 Puc1A 中，是一种主要的丰度蛋白。在高光强下形成的 LH2 复合物中，Puc2A 成分占主导地位，然后在适应低光的 3 至 24 小时内，Puc1B 水平增加了 2.5 倍以上，同时 Puc2A 水平降低了近两倍。这表明 puc1BAC 操纵子受到更严格的光照控制，这可能反映了 puc1 上游调控区的差异。相比之下，在黑暗中低通气下非诱导性地诱导 ICM 形成 48 小时后，Puc2 多肽水平升高，而在适应低光的 24 小时后，上 LH2 富集凝胶带中 Puc 多肽分布没有变化，尽管总体 LH2 水平增加了约两倍。这与该条带源自早期发育过程中形成的 LH2 池一致，该池与随后组装的仅 LH2 结构域不同，形成 LH2 凝胶带。
