Wang Yung-Lin, Ye Li-Ci, Chang San-Chi, Chen Sheng-Chia, Hsu Chun-Hua
Institute of Biochemistry and Molecular Biology, National Yang Ming Chiao Tung University, Taipei 112, Taiwan.
Department of Life Sciences and Institute of Genome Sciences, National Yang Ming Chiao Tung University, Taipei 112, Taiwan.
Int J Biol Macromol. 2025 Jan;284(Pt 1):137999. doi: 10.1016/j.ijbiomac.2024.137999. Epub 2024 Nov 24.
Poly((R)-3-hydroxybutyrate) (PHB) is a microbial biopolymer widely used in commercial biodegradable plastics. PHB degradation in cell is catalyzed by PHB depolymerase (PhaZ), which hydrolyzes the polyester into mono- and/or oligomeric (R)-3-hydroxylbutyrates (3HB). A novel intracellular PhaZ from Bacillus thuringiensis (BtPhaZ) was identified for potential applications in polymer biodegradation and 3HB production. Herein, we present the crystal structure of BtPhaZ at 1.42-Å resolution, making the first crystal structure for an intracellular PhaZ. BtPhaZ comprises a canonical α/β hydrolase catalytic domain and a unique α-helical cap domain. Despite lacking sequence similarity, BtPhaZ shares high structural homology with many α/β hydrolase members, exhibiting a similar active-site architecture. Alongside the most conserved superfamily signature, several new conserved signatures have been identified, contributing not only to the formations of the Ser-His-Asp catalytic triad and the oxyanion hole but also to the active-site conformation. The putative P-1 subsite appears to have limited space for accommodating only one 3HB-monomer, which may provide an explanation why the major hydrolytic product for BtPhaZ is monomeric form. Furthermore, a cluster of solvent-exposed hydrophobic residues in the helical cap domain forms an adsorption site for polymer-binding. Detailed structural comparisons reveal that various PhaZs employ distinct residues for the biopolymer-binding and hydrolysis.
聚（R）-3-羟基丁酸酯（PHB）是一种微生物生物聚合物，广泛应用于商业可生物降解塑料。细胞内的PHB降解由PHB解聚酶（PhaZ）催化，该酶将聚酯水解为单聚和/或低聚的（R）-3-羟基丁酸酯（3HB）。一种来自苏云金芽孢杆菌的新型细胞内PhaZ（BtPhaZ）被鉴定出来，具有在聚合物生物降解和3HB生产中的潜在应用价值。在此，我们展示了分辨率为1.42 Å的BtPhaZ晶体结构，这是首个细胞内PhaZ的晶体结构。BtPhaZ包含一个典型的α/β水解酶催化结构域和一个独特的α-螺旋帽结构域。尽管缺乏序列相似性，但BtPhaZ与许多α/β水解酶成员具有高度的结构同源性，呈现出相似的活性位点结构。除了最保守的超家族特征外，还鉴定出了几个新的保守特征，这些特征不仅有助于丝氨酸-组氨酸-天冬氨酸催化三联体和氧阴离子孔的形成，还对活性位点构象有贡献。推测的P-1亚位点似乎只有有限的空间来容纳一个3HB单体，这可能解释了为什么BtPhaZ的主要水解产物是单体形式。此外，螺旋帽结构域中一组暴露于溶剂的疏水残基形成了一个聚合物结合的吸附位点。详细的结构比较表明，各种PhaZ在生物聚合物结合和水解方面使用不同的残基。
