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Affinity Purification of NF1 Protein-Protein Interactors Identifies Keratins and Neurofibromin Itself as Binding Partners.

Genes (Basel). 2019; 
Carnes RM, Kesterson RA, Korf BR, Mobley JA, Wallis D.
Products/Services Used Details Operation
Catalog Antibody … Cell Signaling Technologies Cat #: 14623S; Danvers, MA, USA), the His tag antibody (GenScript Cat #: A00186; Nanjing, China), or the Halo tag antibody (Promega Cat #: G921A; Madison, WI, USA) The next morning, the … Get A Quote

摘要

Neurofibromatosis Type 1 (NF1) is caused by pathogenic variants in the NF1 gene encoding neurofibromin. Definition of NF1 protein-protein interactions (PPIs) has been difficult and lacks replication, making it challenging to define binding partners that modulate its function. We created a novel tandem affinity purification (TAP) tag cloned in frame to the 3' end of the full-length murine Nf1 cDNA (mNf1). We show that this cDNA is functional and expresses neurofibromin, His-Tag, and can correct p-ERK/ERK ratios in NF1 null HEK293 cells. We used this affinity tag to purify binding partners with Strep-Tactin®XT beads and subsequently, identified them via mass spectrometry (MS). We found the tagged mNf1 can affini... More

关键词

Ras; affinity purification; binding partners; immunoprecipitation; keratins; mass spectrometry; neurofibromatosis Type I; neurofibromin; protein–protein interactors