Mu transpososome activity-profiling yields hyperactive MuA variants for highly efficient genetic and genome engineering.

摘要: The phage Mu DNA transposition system provides a versatile species non-specific tool for molecular biology, genetic engineering and genome modification applications. is catalyzed by MuA transposase, with cleavage integration reactions ultimately attaching the transposon to target DNA. To improve activity of machinery, we mutagenized protein screened hyperactivity-causing substitutions using an in vivo assay. individual activity-enhancing were mapped onto MuA-DNA complex structure, containing tetramer two end segments This analysis, combined varying effect mutations different assays, implied that exert their effects several ways, including optimizing protein-protein protein-DNA contacts. Based on these insights, engineered highly hyperactive versions MuA, combining synergistically acting located subdomains protein. Purified variants are now ready use as second-generation tools variety Mu-based These will also widen scope gene transfer technologies toward medical applications such human therapy. Moreover, work platform further design custom transposases.