The NTAQ1 Knockout HEK293T Cell Line is a CRISPR/Cas9-edited knockout cell line derived from human embryonic kidney HEK293T cells, designed for loss-of-function studies of the NTAQ1 gene (Homo sapiens). This model disrupts the N-terminal glutamine amidase NTAQ1, providing a clean genetic background to dissect the N-end rule pathway of protein degradation. The knockout is generated via CRISPR/Cas9-mediated gene disruption, creating a stable cell line suitable for routine transfection and downstream biochemical assays. It is supplied as a live cell line for basic research applications in cell biology and protein homeostasis.
HEK293T cells are a widely used derivative that stably expresses the SV40 large T antigen, promoting episomal replication and enabling high-level transient protein expression along with efficient production of lentiviral and retroviral vectors. Derived from human embryonic kidney, these cells offer a relevant cellular context for studying protein synthesis, folding, and degradation, particularly the ubiquitin-proteasome system. Their exceptional transfectability and robust recombinant protein output make them a preferred host for mechanistic dissection of protein turnover pathways and functional assays.
NTAQ1 encodes N-terminal glutamine amidase, which deamidates N-terminal glutamine to glutamate on protein substrates, initiating the N-end rule pathway. This creates an N-degron arginylated by ATE1, recognized by UBR1/UBR2 E3 ligases, and degraded by the 26S proteasome. NTAQ1 functions upstream of arginylation and ubiquitination, coupling N-terminal modification to proteasomal turnover. Its substrates include proteins with exposed N-terminal glutamine, which are processed through the ATE1-UBR-proteasome axis after deamidation.
In the HEK293T background, NTAQ1 knockout permits precise evaluation of how N-terminal glutamine deamidation influences protein stability. The line is well-suited for N-degron reporter assays, leveraging HEK293T??s high transfection efficiency to quantify substrate stabilization. Loss of NTAQ1 stabilizes proteins normally targeted for degradation, enabling measurement of half-life changes via cycloheximide chase experiments. This model also facilitates rescue experiments with NTAQ1 variants to probe structure-function relationships within the N-end rule pathway.
Typical applications include cycloheximide chase assays for protein turnover, N-degron reporter assays, and proteasome inhibition experiments. Mass spectrometry-based N-terminomics can identify novel substrates by comparing N-terminal modification profiles between knockout and wild-type cells. Western blotting and RT-qPCR confirm gene disruption and downstream effects. This knockout line also supports drug discovery screens for modulators of the N-end rule pathway. For further information, please contact Ascent Research.
