School of Biological Sciences, Nanyang Technological University, Singapore, 637551, Singapore.
Sci Rep. 2017 Jul 21;7(1):6140. doi: 10.1038/s41598-017-06598-x.
Hyperdisulfide-constrained peptides are distinguished by their high stability and diverse functions. Thus far, these peptides have been reported from animals only but their occurrence in plants are rare. Here, we report the discovery, synthesis and characterization of a hyperdisulfide-constrained peptides family of approximately 2 kDa, β-ginkgotides (β-gB1 and β-gB2) from Ginkgo biloba. Proteomic analysis showed β-ginkgotides contain 18‒20 amino acids, of which 16 residues form a conserved six-cysteine core with a highly clustered cysteine spacing of C‒CC‒C‒CC, an arrangement that has not been reported in cysteine-rich peptides. Disulfide mapping revealed a novel disulfide connectivity of CysI‒IV, CysII‒VI and CysIII‒V. Oxidative folding of synthetic β-gB1 to the native form was obtained in 70% yield. The synthetic β-gB1 displays a compact structure with no regular secondary structural elements, as determined by NMR spectroscopy. Transcriptomic analysis showed precursor βgb1 has a four-domain architecture and revealed an additional 76 β-ginkgotide-like peptides in 59 different gymnosperms, but none in angiosperms. Phylogenetic clustering analysis demonstrated β-ginkgotides belong to a new cysteine-rich peptide family. β-Ginkgotide is resistant to thermal, chemical and proteolytic degradation. Together, β-ginkgotides represent the first-in-class hyperdisulfide-constrained peptide family from plants with a novel scaffold that could be useful for engineering metabolically stable peptidyl therapeutics.
富含二硫键的肽因其高度稳定性和多样化的功能而备受关注。迄今为止，这些肽仅在动物中被报道过，而在植物中则很少见。在这里，我们报告了从银杏中发现、合成和表征一种富含二硫键的肽家族，约 2 kDa，β-银杏肽（β-gB1 和 β-gB2）。蛋白质组学分析表明，β-银杏肽含有 18-20 个氨基酸，其中 16 个残基形成一个保守的六半胱氨酸核心，具有高度聚集的半胱氨酸间隔 C‒CC‒C‒CC，这种排列在富含半胱氨酸的肽中尚未报道过。二硫键图谱显示了 CysI‒IV、CysII‒VI 和 CysIII‒V 的一种新的二硫键连接。通过氧化折叠，合成的 β-gB1 以 70%的收率转化为天然形式。NMR 光谱表明，合成的 β-gB1 具有紧凑的结构，没有规则的二级结构元件。转录组分析表明，前体 βgb1 具有四结构域结构，并在 59 种不同的裸子植物中发现了另外 76 种β-银杏肽样肽，但在被子植物中没有发现。系统发育聚类分析表明，β-银杏肽属于一种新的富含半胱氨酸的肽家族。β-银杏肽能耐受热、化学和蛋白水解降解。总之，β-银杏肽代表了植物中第一个富含二硫键的肽家族，具有一种新的支架，可用于工程代谢稳定的肽类治疗药物。
