Schafheutle M E, Setliková E, Timmins P A, Johner H, Gutgesell P, Setlik I, Welte W
Institut für Biophysik und Strahlenbiologie, Freiburg, FRG.
Biochemistry. 1990 Feb 6;29(5):1216-25. doi: 10.1021/bi00457a018.
An active photosystem I (PSI) complex was isolated from the thermophilic cyanobacterium Synechococcus elongatus by a procedure consisting of three steps: First, extraction of photosystem II from the thylakoids by a sulfobetaine detergent yields PSI-enriched membranes. Second, the latter are treated with Triton X-100 to extract PSI particles, which are further purified by preparative isoelectric focusing. Third, anion-exchange chromatography is used to remove contaminating phycobilisome polypeptides. The purified particles show three major bands in sodium dodecyl sulfate gel electrophoresis of apparent molecular mass of 110, 15, and 10 kDa. Charge separation was monitored by the kinetics of flash-induced absorption changes at 820 nm. A chlorophyll/P700 ratio of 60 was found. When the particles are stored at 4 degrees C, charge separation was stable for weeks. The molecular mass of the PSI particles, determined by measurement of zero-angle neutron scattering intensity, was 217,000 Da. The PSI particles thus consist of one heterodimer of the 60-80-kDa polypeptides and presumably one copy of the 15- and 10-kDa polypeptides, respectively.
通过一个包含三个步骤的程序，从嗜热蓝细菌聚球藻中分离出了活性光系统I（PSI）复合物：首先，用一种磺基甜菜碱去污剂从类囊体中提取光系统II，得到富含PSI的膜。其次，用Triton X-100处理后者以提取PSI颗粒，通过制备性等电聚焦进一步纯化。第三，使用阴离子交换色谱法去除污染的藻胆体多肽。纯化后的颗粒在十二烷基硫酸钠凝胶电泳中显示出三条主要条带，表观分子量分别为110、15和10 kDa。通过820 nm处闪光诱导吸收变化的动力学监测电荷分离。发现叶绿素/P700比率为60。当颗粒保存在4℃时，电荷分离在数周内保持稳定。通过测量零角度中子散射强度确定的PSI颗粒的分子量为217,000 Da。因此，PSI颗粒分别由一个60 - 80 kDa多肽的异二聚体和大概一个15 kDa及10 kDa多肽的拷贝组成。
