Size-exclusion chromatography with post-column nucleic acid staining and fluorescence detection for sensitive ribonucleic acid analysis.

RNA-based therapeutics, such as vaccines or gene therapy applications, are steadily gaining in popularity and require extensive analytical characterization with several orthogonal methods to ensure highest purity for safe use. In this study, size-exclusion chromatography (SEC) with multiple detectors was used for characterizing single- and double-stranded (ss/ds) RNA ladders, providing insights into how conformation and viscosity affect the different elution profiles observed for both species in SEC. Furthermore, high-resolution chromatographic separation was merged with the high-sensitivity detection provided by fluorescent dyes by developing an innovative approach relying on SEC with post-column nucleic acid staining and fluorescence detection (SEC-pcs-FLD).

Cryo-EM structures of holo condensin reveal a subunit flip-flop mechanism.

Complexes containing a pair of structural maintenance of chromosomes (SMC) family proteins are fundamental for the three-dimensional (3D) organization of genomes in all domains of life. The eukaryotic SMC complexes cohesin and condensin are thought to fold interphase and mitotic chromosomes, respectively, into large loop domains, although the underlying molecular mechanisms have remained unknown. We used cryo-EM to investigate the nucleotide-driven reaction cycle of condensin from the budding yeast Saccharomyces cerevisiae.

Structural Basis of an Asymmetric Condensin ATPase Cycle.

The condensin protein complex plays a key role in the structural organization of genomes. How the ATPase activity of its SMC subunits drives large-scale changes in chromosome topology has remained unknown. Here we reconstruct, at near-atomic resolution, the sequence of events that take place during the condensin ATPase cycle.

Structural Basis for a Safety-Belt Mechanism That Anchors Condensin to Chromosomes.

Condensin protein complexes coordinate the formation of mitotic chromosomes and thereby ensure the successful segregation of replicated genomes. Insights into how condensin complexes bind to chromosomes and alter their topology are essential for understanding the molecular principles behind the large-scale chromatin rearrangements that take place during cell divisions. Here, we identify a direct DNA-binding site in the eukaryotic condensin complex, which is formed by its Ycg1 HEAT-repeat and Brn1 kleisin subunits.