2'-Substituted Nucleotides for Stable DNA-Encoded Library Tag Ligation
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Solution Overview
Problem
Current methods for synthesizing and deconvoluting large or complex DNA-encoded libraries face challenges in achieving high yields of tag ligation, particularly under diverse reaction conditions, and require stable nucleotide constructs that can withstand high pH and elevated temperatures, while also simplifying the recognition of tag sequences by DNA- or RNA-dependent polymerases.
Innovation Solution
The use of 2′-substituted nucleotides, such as 2′-O-methyl or 2′-fluoro nucleotides, in oligonucleotide tags, along with specific enzyme and agent conditions, including RNA and DNA ligases, polyethylene glycol, and multivalent cations, to improve single-stranded ligation and encoding processes, allowing for more robust screening and identification of small compounds.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If standard nucleotide constructs are used in DNA-encoded libraries, then the library can be synthesized and screened, but the tag ligation yields are insufficient under diverse reaction conditions including high pH and elevated temperature
Solution Approach 1:
The patent modifies the chemical structure of nucleotides by introducing 2'-substitutions (2'-O-methyl, 2'-fluoro, 2'-amino) to enhance the stability of phosphodiester bonds under diverse reaction conditions including high pH and elevated temperature, thereby improving both reliability and productivity of tag ligation
Solution Approach 2:
The patent creates composite nucleotide structures by combining modified sugar moieties (2'-substituted ribose) with standard or modified bases, resulting in hybrid nucleotide constructs that exhibit enhanced thermal and chemical stability while maintaining ligability
2Adaptability or versatility
If complex multi-step synthesis methods are used to create large DNA-encoded libraries, then library diversity increases, but the deconvolution and identification of tags becomes more difficult
Solution Approach 1:
The patent introduces template-dependent polymerization as an intermediary process that converts complex tag sequences into readable formats by using DNA polymerases to synthesize complementary strands, enabling straightforward sequencing and identification even in highly diverse libraries
Solution Approach 2:
The patent replaces direct observation or complex analytical methods with template-dependent polymerization and sequencing technologies, allowing automated and accurate identification of tag sequences through biological replication rather than direct chemical analysis
3Ease of manufacture
If standard ligation conditions are used, then the synthesis process is simple, but the yields of tag ligation are low under challenging conditions such as high pH and elevated temperature
Solution Approach 1:
The patent modifies nucleotide parameters (2'-substitutions) to inherently withstand challenging conditions, allowing standard ligation protocols to be used without compromising yield, thus maintaining ease of manufacture while improving productivity
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enhances the stability and efficiency of tag ligation, enabling the creation of robust DNA-encoded libraries that can be effectively screened and identified, even under challenging conditions, thereby improving the drug discovery process.
Implementation Method 1
the use of one or more enzymes; optionally, the inclusion of error-recognition capabilities in the tag design; and/or the use of one or more agents during ligation
Data Source
AI summary
The present invention relates to oligonucleotide-encoded libraries and methods of tagging such libraries. In particular, the methods and oligonucleotides can include one or more 2′-substituted nucleotides, such as 2′-O-methyl or 2′-fluoro nucleotides, and other conditions or reagents to enhance enzyme ligation or one or more chemical functionalities to support chemical ligation.