Dataset Information

Indentification of RecBCD Auxiliary ATP binding

ABSTRACT: The RecBCD helicase initiates double-stranded break repair in bacteria by processively unwinding DNA with a rate approaching ~1,600 bp·second, but the mechanism enabling such a fast rate is unknown. By using crosslinking followed by mass spectrometry, we revealed evidence for the existence of auxiliary binding sites in the RecC subunit. This was validated by equilibrium and time-resolved binding experiments, ensemble, and single-molecule unwinding assays. The essentiality and functionality of these sites are further demonstrated by their impact on the survival of E.coli after exposure to damage-inducing radiation.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Escherichia Coli

SUBMITTER: Oded Kleifeld

LAB HEAD: Arnon Henn

PROVIDER: PXD031540 | Pride | 2022-05-20

REPOSITORIES: Pride

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Publications

Auxiliary ATP binding sites support DNA unwinding by RecBCD.

Zananiri Rani R Mangapuram Venkata Sivasubramanyan S Gaydar Vera V Yahalom Dan D Malik Omri O Rudnizky Sergei S Kleifeld Oded O Kaplan Ariel A Henn Arnon A

Nature communications 20220404 1

The RecBCD helicase initiates double-stranded break repair in bacteria by processively unwinding DNA with a rate approaching ∼1,600 bp·s<sup>-1</sup>, but the mechanism enabling such a fast rate is unknown. Employing a wide range of methodologies - including equilibrium and time-resolved binding experiments, ensemble and single-molecule unwinding assays, and crosslinking followed by mass spectrometry - we reveal the existence of auxiliary binding sites in the RecC subunit, where ATP binds with l ...[more]

PMID: 35379800

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Project description:Induced pluripotent stem cells (iPSCs) are regarded as a central tool to understand human biology in health and disease. Similarly, iPSCs from closely related species should be a central tool to understand human evolution and to identify conserved and variable patterns of iPSC disease models. Here, we have generated human, gorilla, bonobo and cynomolgus monkey iPSCs. We show that these cells are well comparable in their differentiation potential and generally similar to human, cynomolgus and rhesus monkey embryonic stem cells (ESCs). RNA sequencing reveals that expression differences among clones, individuals and stem cell type are all of very similar magnitude within a species. In contrast, expression differences between closely related primate species are three times larger and most genes show significant expression differences among the analysed species. However, pseudogenes differ more than twice as much, suggesting that evolution of expression levels in primate stem cells is rapid, but constrained. These patterns in pluripotent stem cells are comparable to those found in other tissues except testis. Hence, primate iPSCs reveal insights into general primate gene expression evolution and should provide a rich source to identify conserved and species-specific gene expression patterns for cellular phenotypes. Contributors: Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery, Hannover Medical School, 30625 Hannover, Germany Max Planck Institute for Evolutionary Anthropology, Deutscher Platz 6, D-04103 Leipzig, Germany We used expression profiling to characterize five gorilla, two bonobo and three macaque iPS clones as well as three iPS clones from two human individuals, three human embryonic stem (ES) cell lines and three macaque ES cell lines. We generated tagged RNA-Seq libraries from these 19 samples including four technical replicates (23 samples). Over 100 million single end reads were generated on the Illumina platform.

Dataset Information

Indentification of RecBCD Auxiliary ATP binding

Publications

Auxiliary ATP binding sites support DNA unwinding by RecBCD.

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