A study of the overcoming the weak point of a nutritional supplementary food, Spirulina in nutrition

作为一种营养补充食品，螺旋藻（Spirulina sp.，一种蓝藻）含有大量的7-腺苷基钴胺（假维生素B_<12>），对人体无活性。为克服螺旋藻食品在营养上的薄弱环节，对细菌细胞中假维生素B_<12>的生理功能和生物合成的基本信息进行了研究。假维生素B_<12>的生理功能螺旋藻含有大量的假维生素B_<12>和维生素B_<12>依赖的蛋氨酸合成酶的高比活性。利用分子克隆技术克隆了白颡鱼（S. platensis）维生素B_<12>依赖性蛋氨酸合成酶基因。从细胞中纯化螺旋藻基因组DNA。退化引物是从已知的其他蓝藻甲基安非他明的保守序列中指定的。利用引物对纯化的DNA进行PCR扩增。开放阅读框为3549bp。推导出的氨基酸序列与thhodesmium erythraeum EMS 101甲基具有较高的相似性。维生素B_<12>和s-腺苷- l-蛋氨酸的结合基序是完全保守的。这些结果表明，开放阅读框编码了一个metH。假维生素B_<12>可能是高原葡萄蛋氨酸合成酶的辅酶。为评价假维生素B_<12>在细菌细胞中是否转化为真维生素B_<12>，在培养基中加入5,6-二甲基苯并咪唑作为维生素B_<12>低核苷酸片段的前体。前体的加入对细胞生长有明显的抑制作用。

A nutritional supplementary food, Spirulina (Spirulina sp., a cyanobacterium), contains substantial amount of 7-adenynyl cobaminde (pseudovitamin B_<12>), which is inactive for humans.To overcome the weak point of the Spirulina food in nutrition, fundamental information on physiological function and biosynthesis of pseudovitamin B_<12> in the bacterial cells was studied.1. Physiological function of pseudovitamin B_<12>Spirulina platensis contains substantial amount of pseudovitamin B_<12> and high specific activity of vitamin B_<12>-dependent methionine synthase. Molecular cloning of the vitamin B_<12>-dependent methionine synthase gene form S. platensis was conducted. Spirulina genomic DNA was purified from the cell. The degenerate primers were designated from well-conserved sequences of the known other cyanobacterial MetH. Using the primers, purified DNA was amplified by PCR. An open reading frame is 3549bp. The deduced amino add sequence showed high similarity to Thichodesmium erythraeum EMS 101 MetH. The binding motif of vitamin B_<12> and s-adenosyl-L-methioneni are completely conserved. These results indicated that the open reading frame encodes a metH. Pseudovitamin B_<12> may function as the coenzyme of the methionine synthase in S. platensis.2. Effect of the addition of precursor of B_<12>When 5,6-dimethylbenzimidazol as a precursor of the lower nucleotide moiety of vitamin B_<12> was added to the medium in order to evaluate whether pseudovitamin B_<12> is converted to true vitamin B_<12> in the bacterial cells. The addition of the precursor inhibited significant the cell growth.