The right fit: Synthetic nucleotide analogues are widely used to investigate the mechanisms that govern DNA polymerase selectivity—processes that are crucial for the survival of every living organism. The first crystal structures of size‐augmented 4′‐methylated and 4′‐ethylated thymidine triphosphates (TTPs) in complex with a DNA polymerase have been elucidated (picture: superposition of three DNA polymerase structures in complex with TTPs).
Up to now, the direct ligation of two DNA fragments with opposite directions to obtain 3′-3′ or 5′-5′ phosphate ester bonds is still challenging. The only way to obtain DNA oligonucleotides containing a 3′-3′ or 5′-5′ inversion of polarity sites is based on professional DNA chemical synthesis. Herein, we demonstrate a convenient template-directed chemical ligation that enables 3′-3′ and 5′-5′ linkages of two DNA oligonucleotides. This method is based on the assembly of two oligonucleotides on a template in opposite directions through forming antiparallel and parallel duplexes simultaneously, followed by coupling with N-Cyanoimidazole under mild condition. Moreover, on the basis of DNA oligonucleotides with 5′-5′ linkage obtained through our template-directed chemical ligation, we developed a new cDNA display technique for in vitro selection of functional polypeptides.
Das passt: Synthetische Nucleotidanaloga werden zur Untersuchung von Mechanismen verwendet, die die Selektivität von DNA‐Polymerasen steuern und für das Fortbestehen jedes Organismus entscheidend sind. Die ersten Kristallstrukturen von vergrößerten, 4′‐methylierten und ‐ethylierten Thymidintriphosphaten (TTPs) im Komplex mit einer DNA‐Polymerase wurden nun erhalten (Bild: Überlagerung dreier DNA‐Polymerasestrukturen im Komplex mit TTPs).
Finding the right fit: Herein, we report on the development of novel steric probes and present initial insights into their interplay with DNA polymerases. Our findings provide experimental evidence for varied enzyme-substrate interactions that might account for the varied selectivity previously observed.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.