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Chemical composition engineering is one of the most valid approaches to regulate the active sites for improved activity.
Here we use comparative structural analyses, molecular dynamics simulations, and in silico sequence re-engineering via directed sampling by RosettaDesign to identify key molecular determinants of USP7cd activation and successfully engineer USP7cd for improved activity.
To improve some of the catalytic properties of P2Ox from Trametes multicolor, we selected a semi-rational approach of enzyme engineering, saturation mutagenesis of active-site residues and subsequent screening of mutant libraries for improved activity.
Based on these hypotheses, we attempted in this study, to engineer ALDH for improved activity toward 3-HPA and NAD(P)+ via lowering the Km values for both the substrates.
To verify the triplet-motif, two variants of E. coli AdhZ2 DIN and LND were designed and confirmed for improved activity with NADH.
After two rounds of error-prone PCR and backcross with parental DNA, three mutants were identified for improved activity toward pyrene and for the first time a new activity toward chrysene in comparison to the wild type enzyme.
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Additionally, the large number of attachment sites for nitrogen and atomic iron functionalization provide an avenue for improving activity.
Here we use the KE07 enzyme as a model system to reveal those forces, which govern enzymatic evolution and elucidate the key factors for improving activity.
One avenue for improving activity with crXNA molecules would be through the use of alternative backbone chemistries that may be more tolerated by Cas9, or through evolution of Cas9 to accommodate these molecules more efficiently by improving the conformational changes of the complexes.
A docking study of the most active compound 11g revealed key interactions between 11g and Hsp90-Cdc37 complex in which the benzothiazole moiety and the amine chain group were important for improving activity.
A docking study of the most active compound 8k revealed key interactions between 8k and MMP-3 in which the sulfonamide moiety and the dipeptide group were important for improving activity.
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