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Pipeline publication

[…] in 10 mL HEPES/MgCl2 buffer (50 mM HEPES, 5 mM MgCl2 [pH 6.8]), and resuspended in a volume equivalent to 10 mL for a culture harvested at OD600 3.0. Labeling reactions were performed for 1 hr at room temperature, in a final volume of 100 μL HEPES/MgCl2 buffer containing 80 μL cell suspension, 5 mM malPEG, and 10 mM EDTA. Non-labeled control samples had malPEG omitted. Reactions were stopped by addition of DTT to 100 mM final concentration. Cells were isolated by centrifugation (21,000 × g, 5 min) and resuspended in 100 μL SDS sample buffer prior to SDS-PAGE., Sequence alignments were performed using T-Coffee or MUSCLE (http://www.ebi.ac.uk/Tools/msa/). Structure models were generated using I-TASSER (). Ssp2 model was further regularized by 1 ns molecular dynamics in explicit solvent followed by energy minimization (Gromacs 2016.3; ). Structural alignments were performed in PyMol (version 1.8.6.0; Schrodinger)., Quantitative data were analyzed and presented using GraphPad Prism, and pairwise comparison was performed using unpaired t tests (two tailed). All replicates were independent biological replicates., , This work was supported by the Wellcome Trust (grant 104556/Z/14/Z to S.J.C. and grant 097818/Z/11/B to the University of Dundee). We thank Tracy Palmer for technical advice and access to the Keio collection, Birte Hollman for construction of strain BH03, Jean-François Gaucher for assistance with structural prediction, Katharina Trunk for advice and discussion, and the Dundee Imaging Facility for microscopy support., Study Design, G.M., S.J.C.; Ex […]

Pipeline specifications

Software tools I-TASSER, GROMACS, PyMOL