Journal: Bio-protocol

Article Title: TIE-UP-SIN: A Method for Enhanced Identification of Protein–Protein Interactions

doi: 10.21769/BioProtoc.5663

Figure Lengend Snippet: Western blot (A) and silver-stained gel (B) show the purification fractions of the WT control (WT/WT), bait/WT experiment (bait/WT), and bait control (bait/bait), all crosslinked with 0.2% (w/v) FA. The fractions loaded are lysate (L) (1 μL), flowthrough (FT) (1 μL), wash steps 1 (W1) and 3 (W3) (8 μL), eluate (E) (4 μL), and residue (R) (4 μL). The Chameleon ® Due Pre-Stained protein marker was used for molecular weight reference. The position of the signals for PycA (127.72 kDa) and SigA-TS (45.9 kDa) is marked on the right side. PycA is a biotinylated protein and thus is purified unspecifically by the MagStrep beads.

Article Snippet: MagStrep ® Strep-Tactin ® XT beads (IBA Lifesciences, catalog number: 2-5090-010) 14.

Techniques: Western Blot, Staining, Purification, Control, Residue, Marker, Molecular Weight

Journal: bioRxiv

Article Title: Competing forms of protein-protein association and DNA binding exhibited by BrxC from the BREX phage restriction system

doi: 10.64898/2026.04.09.717308

Figure Lengend Snippet: Panel a: Co-expressed BrxB Aci , BrxC Aci 1-551 and strep-tagged PglZ Aci were affinity purified by streptactin chromatography followed by SEC (left trace). All three proteins remain associated through this purification scheme (SDS PAGE, right). Panel b: Mass photometry analyses of the purified BrxB Aci :PglZ Aci dimeric complex (‘B:Z’) incubated with either BrxC Aci 1-551 and ATPψS (gray) or BrxC Aci (E269Q) 1-551 and ATP (orange). The predicted mass of the B:Z complex is 122 kDa and that of the C 1-551 monomer is 66 kDa. Complexes consistent with B:Z:(C 1-551 ) 1 and B:Z:(C 1-551 ) 2 stoichiometries are observed. Panel c: X-ray crystallographic structure of a complex consisting of dimeric BrxC Aci (E269Q) 1-551 , full-length BrxB Aci and PglZ Aci 1-98 . The BrxC-BrxC interface and the BrxB-BrxC interface (boxes and insets) involve comparable residues that bind a single copy of ATP between the corresponding protein subunits.

Article Snippet: Streptactin agarose beads (IBA Lifesciences Inc., cat # 2-1250-002) were prepared by pipetting 12 μl of a 50% slurry into a 0.6 ml Eppendorf tube and centrifuged at 2000 x g for 1 min.

Techniques: Affinity Purification, Chromatography, Purification, SDS Page, Incubation

Journal: bioRxiv

Article Title: Competing forms of protein-protein association and DNA binding exhibited by BrxC from the BREX phage restriction system

doi: 10.64898/2026.04.09.717308

Figure Lengend Snippet: Panel a, left: Co-expression of BrxB Aci , strep-tagged PglZ Aci and full-length BrxC Aci demonstrates co-elution of all three subunits by streptactin affinity purification (WC: Whole cell lysate; Sol: filtered soluble lysate; FT: affinity column flow-through). Panel a, right: The streptacin-eluted B:C:Z complex largely dissociates on SEC into BrxC Aci dimers and the B Aci :Z Aci complex. Panel b: Low resolution cryo-EM analysis of particles corresponding to a complex between a BrxC Aci 1-551 -BrxB Aci fusion (teal and red cartoon elements, respectively) and a separate full-length PglZ Aci subunit (blue cartoon) produces a density envelope that agrees closely with a generated predictive model of a BrxC 1-551 -BrxB:PglZ dimer, with the complex held together via interactions between separate ends of each PglZ subunit. Panel c: ATPase assays of BrxB Aci :PglZ Aci and indicated BrxC Aci constructs, comparing each protein alone, or incubated together at 1 µM concentration.

Article Snippet: Streptactin agarose beads (IBA Lifesciences Inc., cat # 2-1250-002) were prepared by pipetting 12 μl of a 50% slurry into a 0.6 ml Eppendorf tube and centrifuged at 2000 x g for 1 min.

Techniques: Expressing, Co-Elution Assay, Affinity Purification, Affinity Column, Cryo-EM Sample Prep, Generated, Construct, Incubation, Concentration Assay