作　　者： ; ; ;

机构地区： 暨南大学生命科学技术学院

出　　处： 《微生物学报》 2006年第3期401-405,共5页

摘　　要： 从富硒螺旋藻(Se richSpirulina platensis,Se-SP)中分离纯化高纯度的含硒别藻蓝蛋白(Se-containingallophycocyanin,Se-APC)并观察其生化特性。羟基磷灰石和DEAE-52柱层析方法结合制备电泳技术纯化Se-APC;光谱扫描、Native-PAGE、SDS-PAGE和IEF方法鉴定Se-APC生化特性;2,3-DAN荧光光度法检测蛋白质中Se含量。结果发现3种高纯度Se-APC的光谱特征分别与APCI、APCII、APCIII吻合;电泳鉴定它们可能都是(αβ)3,α、β亚基分别为18.3和15.7 kDa,其pI值分别为:4.76、4.85和5.02;3种Se-APC中Se含量分别为316、273和408μg/g,Se-APC经0.5mol/L NaSCN解聚和β-巯基乙醇变性处理后,蛋白质中Se含量依次减低并趋于稳定。结果提示Se-SP中APC可结合Se,APC中Se含量与其分子聚态有关,亚基中含Se量稳定,可能是以共价键方式结合,Se-APC生物活性及硒在蛋白质中的结合位点值得深入研究。 Three Se-containing allophycocyanins （Se-APC） with high purity were purified from Se rich Spirulina platensis （Se-sp.） by hydroxyapatite chromatography, DEAE-52 anion-exchange chromatography and native gel preparative electrophoresis. Their biochemicial properties were explored by spectral scanning and electrophoresis analysis of Native-PAGE, SDS-PAGE and IEF on thin slab gel. Protein molecular weight （MW） of APC aggregation was determined by gel filter on Sephadex G-200 column. Se content of native and denatured Se-APC was detected by 2, 3-DAN fluorocence method. According to visible and fluorescence spectral character, three purified fractions of APC were identified to be APCI, APCII and APCIII. Native-PAGE and SDS-PAGE analysis revealed that they all shaped trimer （αβ）3 of α and βsubunit with molecular mass of 18.3kDa and 15.7kDa, whereas APCI contains γ subunit （about 32kDa） visibly and APCIII maybe contain the linker peptide of Lc （8 - 10kDa）based on their MW to be determined of 130.9,98. land 106.30kDa. IEF detection showed that the pl of Se-APCs was 4.76, 4.85 and 5.02 respectively. Se content of three purified Se-APCs were 316, 273 and 408μg/g, which decreased about 25% after deaggregation treatment by 0.50mol/L NaSCN and decreased more than 50% after denaturation treatment by 2-mercaptoethanol and reached to a steady content of 132μg/g on average. These results indicated that Se incorporation into APC had no influence on function of energy transfer as well as biochemical property of APCs, and Se binding with APCs was highly relevant to its aggregation states whereas Se integrated steadily with its subunits.

关 键 词： 螺旋藻 硒 别藻蓝蛋白 纯化

领　　域： [生物学]