penta mutant sortase a his6  (Addgene inc)

Journal: ACS Nano

Article Title: Balancing the Nanoscale Organization in Multivalent Materials for Functional Inhibition of the Programmed Death-1 Immune Checkpoint

doi: 10.1021/acsnano.3c06552

Figure Lengend Snippet: Production of PD-1-DNA conjugates and functionalization on DNA origami disks. (a) Schematic representation of the DNA disk library used to investigate the influence of valency and pattern of PD-1 on the interaction with target ligands. Each black dot represents the position of PD-1 on the DNA disk. (b) Schematic overview of the experimental workflow used to conjugate PD-1 to ssDNA via (1) sortase-mediated reaction and (2) copper-free click chemistry, and (3) hybridize PD-1 to DNA origami disks. Z = poly histidine tag. (c) Schematic representation of the DNA-PAINT experimental setup used for the characterization of 6LH, 6SH, and 6Lin PD-1 DNA disks. PD-1-DNA conjugates were prepared with a ssDNA handle extended with a DNA-PAINT docking sequence (R1). Each PD-1 was visualized by transient hybridization of the imager strand (Cy3B-labeled R1 imager strand) and the docking strand. (d) DNA-PAINT sum images of 6LH, 6SH, and 6Lin PD-1 DNA disks (top). Cross-sectional histogram analysis was performed for the highlighted areas (white boxes) to measure inter-PD-1 distances (bottom). Scale bars: 15 nm.

Article Snippet: The sequence encoding for sortase A5 pentamutent (SrtA) enzyme was obtained from Addgene (Plasmid #75144).

Techniques: Sequencing, Hybridization, Labeling