Journal: Molecular Systems Biology

Article Title: Enabling high‐throughput biology with flexible open‐source automation

doi: 10.15252/msb.20209942

Figure Lengend Snippet:

Article Snippet: pRSET‐B mCherry , Addgene , Cat #108857.

Techniques: Recombinant, Software, Control

Journal: Molecular Systems Biology

Article Title: Enabling high‐throughput biology with flexible open‐source automation

doi: 10.15252/msb.20209942

Figure Lengend Snippet:

Article Snippet: pRSET‐B YFP , Addgene , Cat #108856.

Techniques: Recombinant, Software, Control

Journal: Methods in enzymology

Article Title: Chemiluminescent sensors for quantitation of the bacterial second messenger cyclic di-GMP

doi: 10.1016/bs.mie.2020.04.004

Figure Lengend Snippet: Overview of BRET-based biosensors. (a) General schematic depicting the function of the tVYN-TmΔ biosensor for in vitro experiments. BRET efficiency decreases in the presence of cyclic di-GMP. (b) Cell phone picture taken of a 96-well plate containing biosensor in each well immediately after substrate addition. Biosensor incubated with only H2O in the first column and with increasing concentrations of cyclic di-GMP in other columns. Adapted with permission from ACS Chemical Biology.

Article Snippet: pRSET b -tVYN-TmΔ plasmid [Addgene ID: 149694] pCOLA-PdeH plasmid [Addgene ID: 79165] S.O.C. medium 1 M IPTG stock in ddH 2 O Lysozyme PMSF Sonic dismembrator Ni-NTA resin Chromatography columns with solid support Amicon Ultra-15 centrifugal filter units (10K MWCO) Prestained protein ladder (~10-250 kDa) Polyacrylamide gel electrophoresis equipment Ammonium persulfate (APS) Tetramethylethylenediamine (TEMED) Coomassie Brilliant Blue G-250 Solutions that can be prepared in advance are listed below, while any solutions that should be freshly prepared are detailed in the procedure. ddH 2 O should be used for any solutions containing water.

Techniques: In Vitro, Incubation

Journal: Methods in enzymology

Article Title: Chemiluminescent sensors for quantitation of the bacterial second messenger cyclic di-GMP

doi: 10.1016/bs.mie.2020.04.004

Figure Lengend Snippet: Characteristics of the tVYN-TmΔ biosensor. (a) C-di-GMP affinity measurements with 300 pM tVYN-TmΔ. Measured Kd is less than 300 pM based on data from 3 replicates depicted as mean +/− SD. (b) Example standard curve generated using the described procedure with 3 nM tVYN-TmΔ. Data are from 6 replicates depicted as mean +/− SD. Adapted with permission from ACS Chemical Biology.

Article Snippet: pRSET b -tVYN-TmΔ plasmid [Addgene ID: 149694] pCOLA-PdeH plasmid [Addgene ID: 79165] S.O.C. medium 1 M IPTG stock in ddH 2 O Lysozyme PMSF Sonic dismembrator Ni-NTA resin Chromatography columns with solid support Amicon Ultra-15 centrifugal filter units (10K MWCO) Prestained protein ladder (~10-250 kDa) Polyacrylamide gel electrophoresis equipment Ammonium persulfate (APS) Tetramethylethylenediamine (TEMED) Coomassie Brilliant Blue G-250 Solutions that can be prepared in advance are listed below, while any solutions that should be freshly prepared are detailed in the procedure. ddH 2 O should be used for any solutions containing water.

Techniques: Generated

Journal: bioRxiv

Article Title: Understanding inhibitor resistance in Mps1 kinase through novel biophysical assays and structures

doi: 10.1101/112086

Figure Lengend Snippet: (A) The overall scheme of the FP assay; the peptide (green bar) is fluorescently labelled (yellow star) (B) FP measurements of WT Mps1 kinase activity (blue) by titrating the Bub1/Bub3 complex. Grey and black circles represent the measurement in the presence of NMS-P715 and Cpd-5, respectively.

Article Snippet: The plasmid containing a construct of the Mps1 kinase domain (residues 519–808) was a gift from Dr. Nicola Burgess-Brown (Addgene plasmid # 38907) [ ].

Techniques: FP Assay, Activity Assay

Journal: bioRxiv

Article Title: Understanding inhibitor resistance in Mps1 kinase through novel biophysical assays and structures

doi: 10.1101/112086

Figure Lengend Snippet: (A, C, E) FP assay for the Mps1 kinase variants in the presence of inhibitors; WT and C604Y/W kinase activities are shown in circles and triangles, respectively. IC 50 values were calculated by titrating Cpd-5 (black) and NMS-P715 (grey). (B, D, F) Binding affinity measurements of the Mps1 variants with inhibitors, measured by MST.

Article Snippet: The plasmid containing a construct of the Mps1 kinase domain (residues 519–808) was a gift from Dr. Nicola Burgess-Brown (Addgene plasmid # 38907) [ ].

Techniques: FP Assay, Binding Assay

Journal: bioRxiv

Article Title: Understanding inhibitor resistance in Mps1 kinase through novel biophysical assays and structures

doi: 10.1101/112086

Figure Lengend Snippet: (A) 2mF o -DF c electron density map of Cpd-5 in Mps1 kinase contoured at 1.0σ (carved at 2.5 Å from the Cpd-5 atoms). (B) Cpd-5 in the ATP binding pocket of Mps1 C604Y; Cpd-5, the side chains of Gln541, Tyr604, and Lys553 as well as residue Gly605 are depicted as sticks; hydrogen bonds are depicted as black dotted lines. Figures were made in CCP4mg . (C) Cpd-5 interactions with Mps1 kinase drawn by the Lidia routine in COOT. (D, E, F) The same as A, B, C for the interaction of Cpd-5 with Mps1 C604W. (G, H, I) The same as A, B, C for the interaction of Cpd-5 with Mps1 C604W. (J, K, L) The same as A, B, C for the interaction of NMS-P715 with Mps1 C604W

Article Snippet: The plasmid containing a construct of the Mps1 kinase domain (residues 519–808) was a gift from Dr. Nicola Burgess-Brown (Addgene plasmid # 38907) [ ].

Techniques: Binding Assay, Residue

Journal: bioRxiv

Article Title: Understanding inhibitor resistance in Mps1 kinase through novel biophysical assays and structures

doi: 10.1101/112086

Figure Lengend Snippet: (A) NMS-P715 and Cpd5 in the ATP binding pocket of Mps1 C604Y; (B) NMS-P715 and Cpd5 in the ATP binding pocket of Mps1 C604W; (C) Reversine and Cpd5 in the ATP binding pocket of Mps1 C604Y.

Article Snippet: The plasmid containing a construct of the Mps1 kinase domain (residues 519–808) was a gift from Dr. Nicola Burgess-Brown (Addgene plasmid # 38907) [ ].

Techniques: Binding Assay

Journal: bioRxiv

Article Title: Understanding inhibitor resistance in Mps1 kinase through novel biophysical assays and structures

doi: 10.1101/112086

Figure Lengend Snippet: (A) Cpd-5 in the ATP binding pocket of Mps1 C604Y/W; the C604W structure is shown with thin lines for comparison. (B) NMS-P715 in the ATP binding pocket of Mps1 C604Y/W.

Article Snippet: The plasmid containing a construct of the Mps1 kinase domain (residues 519–808) was a gift from Dr. Nicola Burgess-Brown (Addgene plasmid # 38907) [ ].

Techniques: Binding Assay, Comparison