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Front Cover: A T7 RNA Polymerase Mutant Enhances the Yield of 5′‐Thienoguanosine‐Initiated RNAs (ChemBioChem 2/2018)
Author(s) -
Lyon Seth,
Gopalan Venkat
Publication year - 2018
Publication title -
chembiochem
Language(s) - English
Resource type - Reports
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201700676
Subject(s) - t7 rna polymerase , rna , transcription (linguistics) , exoribonuclease , rna polymerase , mutant , polymerase , riboswitch , biology , chemistry , enzyme , biochemistry , non coding rna , rnase p , escherichia coli , gene , linguistics , philosophy , bacteriophage
The front cover picture shows a one‐pot multi‐enzyme approach that was developed to enhance the yield and purity of RNAs incorporating 5′‐thienoguanosine ( th G). In vitro transcription has been employed to introduce chemical handles and modifications into RNAs, but is typically constrained by poor yields of the modified RNAs. A two‐pronged solution to this long‐standing problem is depicted here: 1) the use of a P266L mutant of T7 RNA polymerase (PDB ID: 3E2E) decreases abortive transcription and improves the total RNA yield as well as percentage incorporation of th G (a fluorescent guanosine surrogate), and 2) the addition of 5′‐pyrophosphohydrolase (RppH) and exoribonuclease I (Xrn1) degrades GTP‐initiated RNAs. This approach results in a better yield of near‐homogenous 5′‐ th G‐incorporated RNA, and should have broad utility for the synthesis of 5′‐modified RNAs. More information can be found in the communication by S. Lyon and V. Gopalan on page 142 in Issue 2, 2018 (DOI: 10.1002/cbic.201700538).