点击次数：

论文类型：期刊论文

发表时间：2019-04-12

发表刊物：JOURNAL OF BIOLOGICAL CHEMISTRY

收录刊物：PubMed、EI、SCIE

卷号：294

期号：15

页面范围：5774-5783

关键字：chitin; carbohydrate; carbohydrate-binding protein; protein structure; Bombyx mori; structural biology; carbohydrate esterase; chitin deacetylase; chitosan; deacetylation activity; SeMet single-wavelength anomalous diffraction

摘要：Insect chitin deacetylases (CDAs) catalyze the removal of acetyl groups from chitin and modify this polymer during its synthesis and reorganization. CDAs are essential for insect survival and therefore represent promising targets for insecticide development. However, the structural and biochemical characteristics of insect CDAs have remained elusive. Here, we report the crystal structures of two insect CDAs from the silk moth Bombyx mori: BmCDA1, which may function in cuticle modification, and BmCDA8, which may act in modifying peritrophic membranes in the midgut. Both enzymes belong to the carbohydrate esterase 4 (CE4) family. Comparing their overall structures at 1.98-2.4 resolution with those from well-studied microbial CDAs, we found that two unique loop regions in BmCDA1 and BmCDA8 contribute to the distinct architecture of their substrate-binding clefts. These comparisons revealed that both BmCDA1 and BmCDA8 possess a much longer and wider substrate-binding cleft with a very open active site in the center than the microbial CDAs, including VcCDA from Vibrio cholerae and ArCE4A from Arthrobacter species AW19M34-1. Biochemical analyses indicated that BmCDA8 is an active enzyme that requires its substrates to occupy subsites 0, +1, and +2 for catalysis. In contrast, BmCDA1 also required accessory proteins for catalysis. To the best of our knowledge, our work is the first to unveil the structural and biochemical features of insect proteins belonging to the CE4 family.