摘要
用ＰＣＲ的方法克隆出了编码蓝细菌Ｓｙｎｅｃｈｏｃｏｃｃｕｓｓｐ．ＰＣＣ７００２ＦＮＲ中ＦＮＲ区的基因ｐｅｔＨＬ，克隆到达载体ｐＥＴ３ａ上，转化大肠杆菌ＢＬ２１（ＤＥ３）后实现了大量表达。重组ＦＮＲ区（ｒＦＮＲＤ）经ＤＥＡＥＳｅｐｈｄｅｘＡ５０离子交换层析及ＳｅｐｈａｄｅｘＧ１００凝胶层析得到大量的电泳均一的ｒＦＮＲＤ。Ｎ末端氨基酸序列分析表明，表达产物确为ｐｅｔＨＬ所编码。且起始Ｍｅｔ翻译后未被除去。ｒＦＮＲＤ与ｒＦＮＲ的吸收光谱相同，其黄递酶活性的最适ｐＨ和最适温度也相同。
The petHL of Synechnococcus sp. PCC 7002 encoding FNR domain (FNRD) of FNR was amplified by PCR, and cloned into expressing vector pET 3a. Overexpression of petHL was achieved with E. coli BL21 (DE3). The recombinant FNRD was purified to homogeneity by DEAE Sephadex A 50 and Sephadex G 100 chromatography. N terminal amino acid sequencing showed that rFNRD was encoded by petHL and initial Met was not posttranslationally removed. rFNRD had the same absorption spectrum, optimal pH and optimal temperature as those of rFNR. rFNRD could catalyze photosynthetic electron transport from P700 to NADP + in vitro.
基金
国家自然科学基金
