Beale S I, Cornejo J
Division of Biology and Medicine, Brown University, Providence, Rhode Island 02912.
J Biol Chem. 1991 Nov 25;266(33):22333-40.
An enzyme extract from the phycocyanin-containing unicellular rhodophyte, Cyanidium caldarium, reductively transforms biliverdin IX alpha to phycocyanobilin, the chromophore of phycocyanin, in the presence of NADPH. Unpurified cell extract forms both 3(E)-phycocyanobilin, which is identical to the major pigment that is released from phycocyanin by methanolysis, and 3(Z)-phycocyanobilin, which is obtained as a minor methanolysis product. After removal of low molecular weight material from the cell extract, only 3(Z)-phycocyanobilin is formed. 3(E)-Phycocyanobilin formation from biliverdin IX alpha, and the ability to isomerize 3(Z)-phycocyanobilin to 3(E)-phycocyanobilin, are reconstituted by the addition of glutathione to the incubation mixture. Partially purified protein fractions derived from the initial enzyme extract form 3(Z)-phycocyanobilin plus two additional, violet colored bilins, upon incubation with NADPH and biliverdin IX alpha. Further purified protein fractions produce only the violet colored bilins from biliverdin IX alpha. One of these bilins was identified as 3(Z)-phycoerythrobilin by comparative spectrophotometry, reverse-phase high pressure liquid chromatography, and 1H NMR spectroscopy. A C. caldarium protein fraction catalyzes the conversion of 3(Z)-phycoerythrobilin to 3(Z)-phycocyanobilin. This fraction also catalyzes the conversion of 3(E)-phycoerythrobilin to 3(E)-phycocyanobilin. The conversion of phycoerythrobilins to phycocyanobilins requires neither biliverdin nor NADPH. The synthesis of phycoerythrobilin and its conversion to phycocyanobilin by extracts of C. caldarium, a species that does not contain phycoerythrin, indicates that phycoerythrobilin is a biosynthetic precursor to phycocyanobilin. The enzymatic conversion of the ethylidine group from the Z to the E configuration suggests that the E-isomer is the precursor to the protein-bound chromophore.
来自含藻蓝蛋白的单细胞红藻——嗜热栖热菌（Cyanidium caldarium）的一种酶提取物，在存在烟酰胺腺嘌呤二核苷酸磷酸（NADPH）的情况下，可将胆绿素IXα还原转化为藻蓝胆素，即藻蓝蛋白的发色团。未纯化的细胞提取物可形成3（E）-藻蓝胆素（与通过甲醇解从藻蓝蛋白释放的主要色素相同）和3（Z）-藻蓝胆素（作为次要甲醇解产物获得）。从细胞提取物中去除低分子量物质后，仅形成3（Z）-藻蓝胆素。通过向孵育混合物中添加谷胱甘肽，可重建从胆绿素IXα形成3（E）-藻蓝胆素以及将3（Z）-藻蓝胆素异构化为3（E）-藻蓝胆素的能力。与NADPH和胆绿素IXα一起孵育时，源自初始酶提取物的部分纯化蛋白质组分可形成3（Z）-藻蓝胆素以及另外两种紫色的胆素。进一步纯化的蛋白质组分仅从胆绿素IXα产生紫色胆素。通过比较分光光度法、反相高压液相色谱法和1H核磁共振光谱法，其中一种胆素被鉴定为3（Z）-藻红胆素。嗜热栖热菌的一种蛋白质组分催化3（Z）-藻红胆素向3（Z）-藻蓝胆素的转化。该组分还催化3（E）-藻红胆素向3（E）-藻蓝胆素的转化。藻红胆素向藻蓝胆素的转化既不需要胆绿素也不需要NADPH。嗜热栖热菌提取物合成藻红胆素并将其转化为藻蓝胆素，嗜热栖热菌是一种不含藻红蛋白的物种，这表明藻红胆素是藻蓝胆素的生物合成前体。亚乙基从Z构型到E构型的酶促转化表明E-异构体是蛋白质结合发色团的前体。
