SAMHD1 Protein-Protein Interaction Assay

Recombinant SAMHD1 was purified as described (42 (link)) and the pull-down assay was performed as described (14 (link)). SAMHD1 and IKKα or IKKβ were precleared with Dynabeads Protein A for 30 min and then incubated with Dynabeads Protein A and SAMHD1 antibody, IKKα antibody (61294S, Cell Signaling) or IKKβ antibody in cell lysis buffer with 0.375% CHAPS overnight at 4 °C. The beads were washed with a buffer containing 50 mM Tris-HCl PH 8.0, 150 mM NaCl, 1% Triton X-100, and 0.5% sodium deoxycholate. The input and IP samples were analyzed by Western blot.

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Yang H., Espada C.E., Phillips S., Martinez N., Kenney A.D., Yount J.S., Xiong Y, & Wu L. (2023). The host antiviral protein SAMHD1 suppresses NF-κB activation by interacting with the IKK complex during inflammatory responses and viral infection. The Journal of Biological Chemistry, 299(6), 104750.

Publication 2023

IKKα antibody

Antibody Assay Buffer Cell Chaps Ikkα Ikkβ Nacl Protein a Pull Samhd1 Sodium deoxycholate Tris Triton x 100 Western blot

Corresponding Organization :

Other organizations : University of Iowa, Yale University, The Ohio State University

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Variable analysis

independent variables

SAMHD1 antibody
IKKα antibody
IKKβ antibody

dependent variables

Interaction between SAMHD1 and IKKα or IKKβ

control variables

Cell lysis buffer with 0.375% CHAPS
Buffer containing 50 mM Tris-HCl pH 8.0, 150 mM NaCl, 1% Triton X-100, and 0.5% sodium deoxycholate

controls

Positive control: Not specified
Negative control: Not specified

Annotations

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