School of Biological Science and Technology, University of Jinan, Jinan, 250022, China.
Protein Pept Lett. 2023;30(6):506-512. doi: 10.2174/0929866530666230509141807.
Poly (ethylene terephthalate) (PET) is one of the most abundant polyester materials used in daily life and it is also one of the main culprits of environmental pollution. ICCG (F243I/D238C/S283C/Y127G) is an enzyme that performs four modifications on the leaf branch compost keratase (LCC). It shows excellent performance in the hydrolysis of PET and has a great potential in further applications.
Here, we used ICCG to degrade PET particles of various sizes and use the density of attack sites (Γ) and kinetic parameters to evaluate the effect of particle size on enzyme degradation efficiency. We are surprised to observe that there is a certain relationship between K and Γ. In order to further confirm the relationship, we obtained three different enzymes (Y95K, M166S and H218S) by site-directed mutagenesis on the basis of ICCG.
The results confirmed that there was a negative correlation between Km and Γattack. In addition, we also found that increasing the affinity between the enzyme and the substrate does not necessarily lead to the increase of degradation rate.
These findings show that the granulation of PET and the selection of appropriate particle size are helpful to improve its industrial application value. At the same time, additional protein engineering to increase ICCG performance is realistic, but it can't be limited to enhance the affinity between enzyme and substrate.
聚对苯二甲酸乙二醇酯（PET）是日常生活中使用最广泛的聚酯材料之一，也是造成环境污染的主要元凶之一。ICCG（F243I/D238C/S283C/Y127G）是一种在叶支角蛋白酶（LCC）上进行四种修饰的酶。它在 PET 水解方面表现出优异的性能，在进一步的应用中有很大的潜力。
在这里，我们使用 ICCG 来降解各种大小的 PET 颗粒，并使用攻击位点密度（Γ）和动力学参数来评估颗粒大小对酶降解效率的影响。我们惊讶地观察到 K 和 Γ 之间存在一定的关系。为了进一步证实这种关系，我们在 ICCG 的基础上通过定点突变获得了三种不同的酶（Y95K、M166S 和 H218S）。
结果证实 Km 和 Γattack 之间呈负相关。此外，我们还发现，增加酶与底物之间的亲和力不一定会导致降解速率的增加。
这些发现表明，PET 的颗粒化和选择适当的颗粒大小有助于提高其工业应用价值。同时，增加 ICCG 性能的额外蛋白质工程是现实的，但不能仅限于增强酶与底物之间的亲和力。
