核磁共振波谱研究脑钠肽空间结构样品制备

The Samples of BNP was been Maked for Studing the Structure of BNP with NMR

用多维核磁共振波谱（ＮＭＲ）研究脑钠肽（ＢＮＰ）空间结构时，必需制备大量的ＢＮＰ样品。用ｐＧＥＸ—３Ｘ作载体与ＢＮＰ—３８ｃＤＮＡ构建成能高效表达ＧＳＴ—ＮＰ—３８融合蛋白的工程菌，表达出的融合蛋白用珠（ＧＳＴ—ａｇａｒｏｓｅｂｅａｄｓ）分离，动态分离的效果是静态分离的三倍；分离时，由振荡２分钟延至３０分钟，分离量可增加１４倍。纯化ＧＳＴ—ＢＮＰ—３８时，动态洗脱的得率是静态洗脱的５．２倍，在动态体系纯化的得率是在静态体系的１０倍。因此，分离纯化ＧＳＴ—ＢＮＰ—３８时，用珠振荡吸附３０分钟，再用ＧＳＨ振荡洗脱３０分钟，可使其得率提高６０倍以上。这为ＮＭＲ研究ＢＮＰ结构时大量样品制备奠定了重要基础。

When the structure of BNP is been studing by NMR,the BNP samples must be made in larg amount.The engineering bacterium has been constructed with the pGEX-3x as a vector and the BNP-38 cDNA as a foreign DNA that expresses the GST-BNP-38 fusion protien.The fusion protein can be separated with GST-agarose beads.The yeild of separation of fusion protein umder rotation condition is 3 times that under non-rotation condition.When the rotating time was increadsed to 30′,the separating qnentity has been increased by 14 times as against that the time was 2′,When the GST -BNP -38 were purificated ,the eluting amount under rotaion condition 52 times that under non-rotation condition.The purification yeild of fusion protein in a rotating system is 10 times that in a non-rotating system.As a result ,when fusion proteins were absorbed for 30′again under rotation condition,the yeild of purification fusion proteins may be increased by 60 times as against that under non-rotation condition.This has set up an important foundation that preparate samples in larg amount for the research of BNP with NMR.