Journal: Science Advances
Article Title: Discovery of diverse and high-quality mRNA capping enzymes through a language model–enabled platform
doi: 10.1126/sciadv.adt0402
Figure Lengend Snippet: ( A ) Diagram of the in vivo capping activity assay. The construction details of the K1.5/T7 transcriptional system used for the in vivo assays are shown in fig. S15. YFP, yellow fluorescent protein. ( B ) The capping activities of 46 text-mined CEs in CHO cells. The RNAP not fused to a CE served as the negative control (NO_CE, blue bar), and the RNAP fused to the NP868R CE served as the positive control (gray bar and blue dashed line). The relative activities were calculated by assigning the NO_CE mean fluorescence intensity as 1. The bar plot shows the means ± SD of three biological replicates. Sequences chosen for yeast evaluation are marked by orange dots. Data are found in table S5. ( C ) The capping activities of 18 text-mined CEs in S. cerevisiae . Controls, error bars, and color coding are the same as in (B). ( D ) Diagram of the in vitro capping activity assay. ( E ) The in vitro capping activities of two CEs (MRV_5 and MRV_6) found in this work compared with commercial vaccinia CE (VCE) (control_NEB) and no enzyme treatment control (control_NoEnzyme). Each group includes three biological replicates, with representative mass spectra shown. The x axis represents the molecular weight of the compounds, with the observed mass displayed above the corresponding peaks (amu, the relative atomic mass unit). The y axis is the count of compounds collected. For compound identities and liquid chromatography–mass spectrometry (LC-MS) results of all experiments, see figs. S22 to S25 and table S6. The bar plot shows the means ± SD of three biological replicates. P values were determined using an unpaired two-sided t test.
Article Snippet: With the same enzyme usage and reaction conditions, both MRV_5 and MRV_6 [Marseilleviridae CEs (MCEs)] exhibited twofold enhanced capping efficiencies to the commercial vaccinia CE (VCE) from New England Biolabs (NEB; and figs. S22 to S25).
Techniques: In Vivo, Activity Assay, Negative Control, Positive Control, Fluorescence, In Vitro, Control, Molecular Weight, Liquid Chromatography, Mass Spectrometry, Liquid Chromatography with Mass Spectroscopy