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Failure of POM5 binding to the mutated mouse PrP was also confirmed by SPR (data not shown).
Moreover, we confirmed the lack of AP-1 binding to the mutated AP-1 2mut AP-1 2mut3mut probes (dand not shown).
There was a significant decrease in RUNX1 binding to the mutated sequence compared to the common allele (wt-probe) (Fig. 2a, lanes 2 and 4).
However, if the binding of proteins to EGFR inhibits the catalyzation of EGFR ubiquitination and only free EGFR acts as the substrate of Cbl, decreased protein binding to the mutated EGFR would promote efficient EGFR ubiquitination due to the increase in the amount of free EGFR.
These deleterious effects observed with mutant viral stocks are likely to be a direct consequence of impaired AP-1 binding to the mutated 5103 fragment since wild-type and mutant HIV-1 particles from the viral stocks used in the infection studies are structurally similar at both the RNA and protein levels.
The calculated Kd values are 28 µM and 33 µM in the presence of ATP and ADP, respectively (Fig. 4C), starting from a Kd value of 20 µM in absence of ATP or ADP (it should be noted that DAPdel Kd value was 5.5 µM in the 250 mM NH4Ac buffer pH 8.8, suggesting an impact of pH on the probe binding to the mutated protein).
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However, the active metabolite that binds to the mutated ligand-binding domain of the human estrogen receptor is 4-hydroxy-tamoxifen (4-OHT).
This implies that P5 most likely stimulates LNCaP-cell proliferation through binding to the cellular mutated AR present in LNCaP cells.
As shown in Figure 2B, the RNA oligonucleotide comprising the wild-type sequence bound Sam68 in a dose-dependent manner (lanes 1 4), whereas binding to the A/C-mutated version was severely compromised (lanes 5 8), indicating that Sam68 binds to the oligonucleotide depending on this sequence.
Both yeast one-hybrid analysis and co-expression of reporter and effector genes demonstrated that two proteins, TaWRKY and TaRAV, which can specially bind to TaeIF5A1 promoter fragments containing the W-box, but failed in binding to the same fragments containing the mutated core sequence "T TTT".
We found that both TaWRKY and TaRAV can specifically bind to the two promoter fragments containing the W-box, but failed in binding to the promoter fragment containing the mutated core sequence "T TTT" and the control (N1, N2), indicating that they may regulate the expression of TaeIF5A1 through binding to the W-box motif in the promoter of TaeIF5A1.
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