Conformational transitions of Streptococcus pyogenes Cas9 induced by salt and single-guide RNA binding.

Streptococcus pyogenes (Sp) Cas9 has been widely utilized to edit genomes across diverse species. To achieve high efficiency and specificity as a gene editing enzyme, Sp Cas9 undergoes a series of sequential conformational changes during substrate binding and catalysis. Here, we employed single molecule FRET techniques to investigate the effect of different KCl concentrations on conformational dynamics of Sp Cas9 in the presence or absence of a single-guide RNA (sgRNA).

Mechanistic Basis for a Single Amino Acid Residue Mutation Causing Human DNA Ligase 1 Deficiency, A Rare Pediatric Disease.

In mammalian cells, DNA ligase 1 (LIG1) functions as the primary DNA ligase in both genomic replication and single-strand break repair. Several reported mutations in human LIG1, including R305Q, R641L, and R771W, cause LIG1 syndrome, a primary immunodeficiency. While the R641L and R771W mutations, respectively located in the nucleotidyl transferase and oligonucleotide binding domains, have been biochemically characterized and shown to reduce catalytic efficiency, the recently reported R305Q mutation within the DNA binding domain (DBD) remains mechanistically unexplored.