In vitro selection of a 3′ terminal short protector that stabilizes transcripts to improve the translation efficiency in a wheat germ extract. Issue 16 (15th August 2019)

• Record Type: Journal Article
• Authors: Ogawa, Atsushi; Kutsuna, Akane; Takamatsu, Masashi; Okuzono, Tatsuya
• Is Part Of: Bioorganic & medicinal chemistry letters. Volume 29:Issue 16(2019)
• Page Start: 2141
• Page End: 2144
• Publication Date: 2019-08-15
• Subjects: Cell-free translation -- Wheat germ extract -- In vitro selection -- RNA stabilization -- Translational enhancer; Bioorganic chemistry -- Periodicals; Pharmaceutical chemistry -- Periodicals; 572
• DOI: 10.1016/j.bmcl.2019.06.058
• Languages: English
• ISSNs: 0960-894X

Abstract

Graphical abstract: Abstract: Wheat cell-free expression systems based on wheat germ extract (WGE) enable us to briefly synthesize various types of proteins in vitro merely by exogenously adding their mRNA templates. Moreover, it is possible to produce larger amounts of protein by thoroughly removing the endosperm, which contains many translation inhibitors, including ribonucleases (RNases). However, because small amounts of RNases are also present even in an endosperm-free, high-quality WGE (hqWGE), the in-vitro transcribed mRNA is rapidly degraded. In particular, 3′ exonucleases have been considered as the major RNases that degrade mRNA. We thus herein performed in vitro selection to find an effective, short 3′ protector sequence from a random RNA pool. The selected sequences stabilized in vitro transcripts in the hqWGE more effectively than the previously reported, longer 3′ protectors did. In addition, when one of these 3′ protectors was minimized and then fused to mRNA, the translation efficiency increased 5–6-fold in the hqWGE, mainly due to the mRNA stabilization.