多酶共固定化技术能充分发挥不同酶的特点，连续催化多步反应。但固定化后，酶活性中心取向不易控制，酶活性损失显著，将酶进行定向固定化是解决这一问题的有效方法。为此，申请人提出了两种新型的酶定向共固定化方法，亲和LB膜法和共辅基法。亲和LB膜法选择与酶活性中心具有亲和作用的分子，对其改性使其具有表面活性，制备单分子膜。通过亲和作用，酶分子选择性定向结合在单分子膜下。将酶分子转移到载体，得到酶催化活性中心向外的固定化酶。这种方法易于实现多酶体系的定向共固定化，达到连续催化的目的。对于生物传感器，希望酶催化活性中心直接与电极接触。对此，可采用共辅基法，即将几种辅基共固定化于一个导电高分子上，通过控制导电高分子的分子量精确调控酶分子间的距离，实现多酶体系协同效应。本项目为设计制备高活性定向固定化多酶体系提供了理论依据，对DNA和抗体等生物活性大分子固定化也具有重要的借鉴意义。

Multienzyme system can catalyze a wide variety of metabolic processes, which involve multiple reaction steps. After immobilization, enzyme always lost its activity largely. To avoid this drawback, oriented immobilization of enzyme is regarded as an efficient method. In our work, two novel methods of the oriented co-immobilization of multienzyme system with higher enzyme activity are presented, affinity Langmuir-Blodgett and co-prosthetic group. For affinity Langmuir-Blodgett method, a ligand which can form affinity interaction with the activity site of enzyme, is modified with a long carbon chain. Then, the modified ligand is used to prepare a single molecular film on the water/air surface. After that, the enzyme form affinity interaction with the film. Last, the enzyme is transferred to support to obtain the multienzyme system of oriented co-immobilization. For co-prosthetic group method, which is efficient for biosensor to directly contact the active site of enzyme with electrode, the molecular structure of conductive polymer is designed to co-immobilize prosthetic group, and the molecular weight of this conductive polymer is control to tailor the distance of enzymes. This work would be helpful to provide theory found for the oriented co-immobilization of multienzyme system with higher enzyme activity and biofunctional materials containing other biomolecules (sucha as DNA and antibody).