Liu Y C, Ledger R, Stellwagen E
J Biol Chem. 1984 Mar 25;259(6):3796-9.
The interaction of rabbit muscle lactate dehydrogenase with reactive blue 2 immobilized on Sepharose CL-6B was quantitatively evaluated by analytical procedures using both frontal and zonal chromatography and also static equilibrium methodology. All three analytical procedures give a common result, namely (i) that each enzyme molecule is retained by a single immobilized dye molecule; (ii) that the immobilized dye binds to the protein at an NADH binding site with an affinity identical to that of the free dye; and (iii) that less than 2% of the immobilized dye is accessible to the enzyme. The very small fractional accessibility is not due to steric exclusion but likely is a result of adsorption of the immobilized dye to the matrix surface.
采用前沿色谱法、区带色谱法和静态平衡法等分析方法，对固定在琼脂糖凝胶CL - 6B上的活性蓝2与兔肌肉乳酸脱氢酶之间的相互作用进行了定量评估。所有这三种分析方法都得出了相同的结果，即：（i）每个酶分子被单个固定化染料分子保留；（ii）固定化染料在NADH结合位点与蛋白质结合，其亲和力与游离染料相同；（iii）酶可接触到的固定化染料不到2%。如此小的可及分数并非由于空间排斥，而可能是固定化染料吸附到基质表面的结果。
