Genetic Code Expansion of the Silkworm Bombyx mori Using a Pyrrolysyl-tRNA Synthetase/tRNA^Pyl Pair.

IF 3.9 2区生物学 Q1 BIOCHEMICAL RESEARCH METHODS ACS Synthetic Biology Pub Date : 2025-01-17 Epub Date: 2024-12-16 DOI:10.1021/acssynbio.4c00684

Hidetoshi Teramoto, Katsura Kojima

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Abstract

The domesticated silkworm Bombyx mori, an essential industrial animal for silk production, has attracted attention as a host for protein production due to its remarkable protein synthesis capability. Here, we applied genetic code expansion (GCE) using a versatile pyrrolysyl-tRNA synthetase (PylRS)/tRNA^Pyl pair from Methanosarcina mazei to B. mori; GCE enables synthetic amino acid incorporation into proteins to give them non-natural functions. Transgenic B. mori lines expressing M. mazei PylRS and its cognate tRNA^Pyl were generated and cross-mated to obtain their F₁ hybrid. Orally administering a click-compatible synthetic amino acid, trans-cyclooctene-lysine (TCO-Lys), to the F₁ hybrid has led to the production of silk fiber incorporated with TCO-Lys. TCO-Lys incorporation in silk fiber was verified by selective labeling of the TCO group by click chemistry. The developed system is available for large-scale protein production with a wide variety of synthetic amino acids.

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利用pyrolyyl - trna合成酶/tRNAPyl对扩增家蚕遗传密码。

家蚕是一种重要的工业动物，由于其卓越的蛋白质合成能力，作为蛋白质生产的宿主而受到关注。本研究利用多用途pyrlyyl - trna合成酶(PylRS)/tRNAPyl对mazei Methanosarcina mori的遗传密码扩增（GCE）；GCE使合成氨基酸与蛋白质结合，赋予它们非天然的功能。产生了表达mazei PylRS及其同源tRNAPyl的转基因家蚕系，并进行了杂交，获得了其F1杂种。口服一种与点击兼容的合成氨基酸，反式环烯赖氨酸（TCO-Lys），导致F1杂交产生含有TCO-Lys的丝纤维。通过点击化学对TCO基团的选择性标记，验证了TCO- lys在真丝纤维中的掺入。该系统可用于多种合成氨基酸的大规模蛋白质生产。

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来源期刊

ACS Synthetic Biology 生物-

CiteScore

8.00

自引率

10.60%

发文量

380

审稿时长

6-12 weeks

期刊介绍： The journal is particularly interested in studies on the design and synthesis of new genetic circuits and gene products; computational methods in the design of systems; and integrative applied approaches to understanding disease and metabolism. Topics may include, but are not limited to: Design and optimization of genetic systems Genetic circuit design and their principles for their organization into programs Computational methods to aid the design of genetic systems Experimental methods to quantify genetic parts, circuits, and metabolic fluxes Genetic parts libraries: their creation, analysis, and ontological representation Protein engineering including computational design Metabolic engineering and cellular manufacturing, including biomass conversion Natural product access, engineering, and production Creative and innovative applications of cellular programming Medical applications, tissue engineering, and the programming of therapeutic cells Minimal cell design and construction Genomics and genome replacement strategies Viral engineering Automated and robotic assembly platforms for synthetic biology DNA synthesis methodologies Metagenomics and synthetic metagenomic analysis Bioinformatics applied to gene discovery, chemoinformatics, and pathway construction Gene optimization Methods for genome-scale measurements of transcription and metabolomics Systems biology and methods to integrate multiple data sources in vitro and cell-free synthetic biology and molecular programming Nucleic acid engineering.