钝顶螺旋藻藻胆蛋白的分离、纯化及其理化特性

PURIFICATION AND PHYSIOCHEMICAL PROPERTES OF PHYCOBILIPROTEIN OF SPIRULINA PLATENSIS VAR. NANJINGENSIS

钝顶螺旋藻(Spirulina Platensis var.nanjingensis)一变异株的水溶性色素精提物,经固体硫酸铵沉淀,羟基磷灰石(HA)和Sephadex G-100柱层析后可分离、纯化出藻蓝蛋白(C-PC)和别藻蛋白(APC)。它们的纯度可分别达到AS 620/A_(277)=4.71;A_(650)/A_(270)=5.62。纯化后的C—PC和APC在聚丙烯酰胺凝胶电泳(PAGE)中仅见一条色带,其最大吸收峰分别在620nm和050nm。经12％的十二烷基硫酸钠—聚丙烯酰胺凝胶电泳(SDS—PAGE),以及高效液相色谱(HPLC)分离,C—PC和APC均可分为α和β两个亚单位。两者的亚单位分子量分别为:C—PC—α,15000;C—PC—β,14500;APC—α,15000;APC—β,13500。依此推算,该藻的C—PC和APC的最小分子量应为29.5kD和28.5kD。经等电电泳法测定,其C—PC和APC的等电点分别在4.8和4.9。氨基酸组成和含量分析结果表明,除色氨酸(Try)未测外,c—PC含有14种氨基酸,APC含有15种氨基酸,两者都缺乏组氨酸(His)和脯氨酸(Pro),C—PC还缺少蛋氨酸(Met)。

Two types of phycobiliproteins, C-phycocyanin (C-PC)and allophycocyanin (APC)were separated just after crude phycobiliproteins from Spirulina platensis var. nanjingensis were precipitated in 50%(W/V)solid ammonium sulphated and then chromatographed on a hydroxylapatite(HA)column, further purified by Sephadex G-100. The C-PC with a purity of 4. 71(A52o/A277) possessed absorption maximum at 620nm, while the APC with a purity of 5. 62(A55o/A27s) had an absorption ? maximum at 650nm. When the purified C-PC and PAC were electrophoresed under nondenaturing polyacryiamide gel electrophoresis(PAGE) condition, only one band was observed (data not shown). Under denaturation (by means of 12% SDS-polyacrylamide slab gels),C-PC yielded two coloured polypeptides with the molecular weight of 15. OkD and 14. 5kD for its subunits,respectively. The C-PC contained 14 amino acids,APC had 15 amino acids. They were absent of histidine(His) and proline (Pro),C-PC simultaneously lacked methionine(Met). The isoeldectric points of C-PC and APC were 4. 8 and 4. 9, respectively.