Genetically-encoded discovery of proteolytically stable bicyclic inhibitors for morphogen NODAL.
Genetically-encoded discovery of proteolytically stable bicyclic inhibitors for morphogen NODAL.
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DOI：
10.1039/d1sc01916c
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发表时间：
2021-07-21
期刊：
影响因子：
8.4
通讯作者：
Derda R
中科院分区：
文献类型：
--
作者：
Wong JY;Mukherjee R;Miao J;Bilyk O;Triana V;Miskolzie M;Henninot A;Dwyer JJ;Kharchenko S;Iampolska A;Volochnyuk DM;Lin YS;Postovit LM;Derda R
在这篇手稿中，我们开发了一种双重对称关键蛋白(TSL)，它将由20种常见氨基酸组成的现成的噬菌体展示肽库转化为展示在噬菌体上的遗传编码的双环肽文库。TSL结合了一个醛反应基团和两个硫醇反应基团；它将半胱氨酸[C]的两个侧链与N-末端丝氨酸衍生的N-末端醛[S]连接起来，得到了一个新型的缺少N-末端的双环拓扑结构。以45±15%的产率将SX1CX2X3X4X5X6X7C序列的噬菌体展示文库转化为双环TSL-[S]X1[C]X2X3X4X5X6X7[C]多肽文库。以该文库和蛋白质形态原结点为靶点，我们发现了在癌细胞中特异性拮抗结点诱导的信号转导的双环大环。在10μM浓度下，两个发现的双环肽可完全抑制结节诱导的P19胚胎癌细胞中Smad2的磷酸化。在10μM浓度下，TSL-[S]Y[C]KRAHKN[C]BYBLE抑制结节诱导的卵巢癌细胞增殖，IC50为1μM，对对照TYK-NU细胞的生长无影响。在72小时的检测过程中，TSL自行车在富含血清的细胞培养液中保持稳定。我们进一步观察了21个不同环大小、氨基酸序列和交联剂几何形状的不同双环大环在小鼠血清和混合蛋白酶(前驱体酶™)中的一般稳定性。TSL-限制性多肽，以扩展先前报道的由交联半胱氨酸侧链形成的噬菌体展示双环结构的谱系。我们预计，这将有助于发现蛋白水解性稳定的针对各种蛋白质靶点的双环抑制剂。二重对称关键蛋白(TSL)可将现成的噬菌体展示的二硫键多肽库转化为蛋白水解性稳定的双环肽。通过对双环噬菌体文库的筛选，发现了一株结节形态原拮抗剂。
In this manuscript, we developed a two-fold symmetric linchpin (TSL) that converts readily available phage-displayed peptides libraries made of 20 common amino acids to genetically-encoded libraries of bicyclic peptides displayed on phage. TSL combines an aldehyde-reactive group and two thiol-reactive groups; it bridges two side chains of cysteine [C] with an N-terminal aldehyde group derived from the N-terminal serine [S], yielding a novel bicyclic topology that lacks a free N-terminus. Phage display libraries of SX1CX2X3X4X5X6X7C sequences, where X is any amino acid but Cys, were converted to a library of bicyclic TSL-[S]X1[C]X2X3X4X5X6X7[C] peptides in 45 ± 15% yield. Using this library and protein morphogen NODAL as a target, we discovered bicyclic macrocycles that specifically antagonize NODAL-induced signaling in cancer cells. At a 10 μM concentration, two discovered bicyclic peptides completely suppressed NODAL-induced phosphorylation of SMAD2 in P19 embryonic carcinoma cells. The TSL-[S]Y[C]KRAHKN[C] bicycle inhibited NODAL-induced proliferation of NODAL-TYK-nu ovarian carcinoma cells with apparent IC50 of 1 μM. The same bicycle at 10 μM concentration did not affect the growth of the control TYK-nu cells. TSL-bicycles remained stable over the course of the 72 hour-long assays in a serum-rich cell-culture medium. We further observed general stability in mouse serum and in a mixture of proteases (Pronase™) for 21 diverse bicyclic macrocycles of different ring sizes, amino acid sequences, and cross-linker geometries. TSL-constrained peptides to expand the previously reported repertoire of phage-displayed bicyclic architectures formed by cross-linking Cys side chains. We anticipate that it will aid the discovery of proteolytically stable bicyclic inhibitors for a variety of protein targets. A two-fold symmetric linchpin (TSL) converts readily available phage-displayed disulfide peptide libraries to proteolytically stable bicyclic peptides. The bicyclic phage library was screened to discover an antagonist of NODAL morphogen.
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影响因子：
7.3
作者：
Giordanetto, Fabrizio;Kihlberg, Jan
通讯作者：
Kihlberg, Jan
影响因子：
14.8
作者：
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通讯作者：
Winter, Greg
影响因子：
3.2
作者：
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通讯作者：
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针对涉及Cripto-1结合的淋巴结前环路的新的抗鼻单克隆抗体。
影响因子：
5.6
作者：
Focà A;Sanguigno L;Focà G;Strizzi L;Iannitti R;Palumbo R;Hendrix MJ;Leonardi A;Ruvo M;Sandomenico A
通讯作者：
Sandomenico A
影响因子：
--
作者：
Gong Y;Guo Y;Hai Y;Yang H;Liu Y;Yang S;Zhang Z;Ma M;Liu L;Li Z;He Z
通讯作者：
He Z