Departamento de Quimica Fisica. Facultad de Ciencias, Unidad de Excelencia de Quimica Aplicada a Biomedicina y Medioambiente (UEQ), Universidad de Granada, 18071, Granada, Spain.
Department of Biology, Georgia State University, Atlanta, GA, 30303, USA.
Nat Commun. 2021 Jan 15;12(1):380. doi: 10.1038/s41467-020-20630-1.
Glycosidases are phylogenetically widely distributed enzymes that are crucial for the cleavage of glycosidic bonds. Here, we present the exceptional properties of a putative ancestor of bacterial and eukaryotic family-1 glycosidases. The ancestral protein shares the TIM-barrel fold with its modern descendants but displays large regions with greatly enhanced conformational flexibility. Yet, the barrel core remains comparatively rigid and the ancestral glycosidase activity is stable, with an optimum temperature within the experimental range for thermophilic family-1 glycosidases. None of the ∼5500 reported crystallographic structures of ∼1400 modern glycosidases show a bound porphyrin. Remarkably, the ancestral glycosidase binds heme tightly and stoichiometrically at a well-defined buried site. Heme binding rigidifies this TIM-barrel and allosterically enhances catalysis. Our work demonstrates the capability of ancestral protein reconstructions to reveal valuable but unexpected biomolecular features when sampling distant sequence space. The potential of the ancestral glycosidase as a scaffold for custom catalysis and biosensor engineering is discussed.
糖苷酶在系统发生上广泛分布，是糖苷键裂解的关键酶。在这里，我们介绍了细菌和真核家族 1 糖苷酶假定祖先的特殊性质。该祖先蛋白与现代后裔共享 TIM 桶折叠，但显示出具有大大增强的构象灵活性的大片段区域。然而，桶核心仍然相对刚性，并且祖先糖苷酶活性稳定，在嗜热家族 1 糖苷酶的实验范围内具有最佳温度。在约 1400 个现代糖苷酶的约 5500 个报告的晶体结构中，没有一个显示结合的卟啉。值得注意的是，祖先糖苷酶紧密结合并以确定的埋藏位置结合血红素。血红素结合使 TIM 桶刚性化，并别构增强催化作用。我们的工作表明，当在遥远的序列空间中采样时，祖先蛋白重建能够揭示有价值但出乎意料的生物分子特征的能力。还讨论了作为定制催化和生物传感器工程支架的祖先糖苷酶的潜力。
