Journal: Molecular cell
Article Title: In vivo evidence for ATPase-dependent DNA translocation by the Bacillus subtilis SMC condensin complex
doi: 10.1016/j.molcel.2018.07.006
Figure Lengend Snippet: A, Homology model of the ATPase domain of B. subtilis SMC. Only one of the two composite ATP binding domains is shown. Amino acids that were mutated are indicated (yellow). A schematic of the condensin complex with its head domains (boxed) bound to ATP (blue balls) is shown on the left. B, Growth of wild-type and the relevant mutants in the presence and absence of ParB on agar plates. The 10−2 and 10−5 dilutions are shown. Under Hi-C assay conditions (22˚C no IPTG and 37˚C with IPTG in CH medium), the mutants grow similar to wild-type. C, NADH-coupled ATPase activity assay for the indicated mutants. D, Bar graph showing ATPase activities. Error bars show the standard deviation of four replicates. See also Figures S1, S2, Tables S1 and S2.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Anti-SMC polyclonal rabbit antibody ( Lindow et al., 2002 ) N/A Anti-ParB polyclonal rabbit antibody ( Lin et al., 1997 ) N/A Anti-ScpA polyclonal rabbit antibody, affinity purified ( Wang et al., 2017 ) N/A Anti-ScpA polyclonal rabbit antibody, affinity purified ( Wang et al., 2017 ) N/A Anti-SigA polyclonal rabbit antibody ( Fujita, 2000 ) N/A Chemicals, Peptides, and Recombinant Proteins Benzonase EMD Cat # 7046 Ni-NTA agarose Qiagen Cat # 30230 HiTrap Q HP GE Healthcare Cat # 17115301 Bradford Assay Kit ThermoFisher Cat # 23236 ATP Dot Scientific Cat # DSA30030–5 NADH Acros Organic Cat # 271100010 Phosphoenolypyruvate Tokyo Chemical Industry Cat # P0256 Rabbit Pyruvate Kinase Roche Diagnostics Cat # 10109045001 Lactate Dehydrogenase Sigma, Darmstadt, Germany Cat # L2500–25KU Formaldehyde 37% Sigma Cat # F8775 Ready-Lyse Lysozyme Epicentre Cat # R1802M HindIII NEB Cat # R0104M Klenow NEB Cat # M0210L Biotin-14-dATP ThermoFisher Cat # 19524016 T4 DNA ligase NEB Cat # M0202M Proteinase K NEB Cat # P8107S T4 DNA Polymerase NEB Cat # M0203L Critical Commercial Assays ATPase/GTPase Activity Assay Kit Sigma Cat # MAK113 NEBNext Ultra II DNA Library Prep Kit NEB Cat # E7645S Deposited Data Raw and analyzed Hi-C data This paper GEO: {"type":"entrez-geo","attrs":{"text":"GSE95137","term_id":"95137"}} GSE95137 Original Data Mendeley Data http://dx.doi.org/10.17632/tygwp234gr.1 Experimental Models: Organisms/Strains Bacillus subtilis strains, see Table S1 Oligonucleotides See Table S3 Recombinant DNA See Table S2 Software and Algorithms MATLAB 8.5 (R2015) ( Wang et al., 2017 ) https://www.mathworks.com/ hiclib ( Imakaev et al., 2012 ) https://bitbucket.org/mirnylab/hiclib Open in a separate window KEY RESOURCES TABLE
Techniques: Binding Assay, Hi-C, Activity Assay, Standard Deviation
