Journal
ACS SYNTHETIC BIOLOGY
Volume 10, Issue 7, Pages 1750-1760Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acssynbio.1c00142
Keywords
TdT polymerase; microarray; synthetic biology; L-DNA; enzymatic DNA synthesis; photolithographic synthesis
Categories
Funding
- Austrian Science Fund [FWF P30596]
- Faculty of Chemistry of the University of Vienna
Ask authors/readers for more resources
The untemplated activity of terminal deoxynucleotidyl transferase (TdT) is influenced by the nature of the initiating strands, including their length, chemistry, and nucleotide composition. Experimental results show that TdT is also catalytically active on a non-nucleosidic substrate, hexaethylene glycol. These findings provide new insights into constraints and strategies for de novo synthesis of DNA using TdT.
The untemplated activity of terminal deoxynucleotidyl transferase (TdT) represents its most appealing feature. Its use is well established in applications aiming for extension of a DNA initiator strand, but a more recent focus points to its potential in enzymatic de novo synthesis of DNA. Whereas its low substrate specificity for nucleoside triphosphates has been studied extensively, here we interrogate how the activity of TdT is modulated by the nature of the initiating strands, in particular their length, chemistry, and nucleotide composition. Investigation of full permutational libraries of mono- to pentamers of D-DNA, L-DNA, and 2'O-methyl-RNA of differing directionality immobilized to glass surfaces, and generated via photolithographic in situ synthesis, shows that the efficiency of extension strongly depends on the nucleobase sequence. We also show TdT being catalytically active on a non-nucleosidic substrate, hexaethylene glycol. These results offer new perspectives on constraints and strategies for de novo synthesis of DNA using TdT regarding the requirements for initiation of enzymatic generation of DNA.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available