The present disclosure relates to compositions, kits, and methods of making RNA vaccines having an appropriate cap structure. Systems, apparatus, compositions, and/or methods may include and/or use, in some embodiments, non-naturally occurring single-chain RNA capping enzymes. In some embodiments, an RNA capping enzyme may include an FCE variant having (a) an amino acid sequence at least 90% identical to positions 1 to 878 of SEQ ID NO: 1, and/or (b) one or more substitutions relative to SEQ ID NO: 1 at a position selected from positions corresponding to positions 215, 337, 572, 648, and 833 (e.g., a position selected from positions corresponding to position 215, 337, and 572) of SEQ ID NO: 1.

Provided herein is a method for efficiently capping RNA in vitro. In some embodiments the capping reaction may be done at high temperature using Vaccinia capping enzyme or a variant thereof. In other embodiments, the capping reactions may comprise a capping enzyme from a large virus of amoeba, e.g., Faustovirus, mimivirus or moumouvirus, or a variant thereof. Compositions and kits for practicing the method are also provided.

The present disclosure relates to compositions, kits, and methods of making RNA vaccines having an appropriate cap structure. Systems, apparatus, compositions, and/or methods may include and/or use, in some embodiments, non-naturally occurring single-chain RNA capping enzymes. In some embodiments, an RNA capping enzyme may include an FCE variant having (a) an amino acid sequence at least 90% identical to positions 1 to 878 of SEQ ID NO: 1, and/or (b) one or more substitutions relative to SEQ ID NO: 1 at a position selected from positions corresponding to positions 215, 337, 572, 648, and 833 (e.g., a position selected from positions corresponding to position 215, 337, and 572) of SEQ ID NO: 1.

The present invention relates to a chimeric enzyme comprising or consisting of at least one catalytic domain of a capping enzyme and at least one RNA-binding domain of a protein-RNA tethering system as well as its application for the production of an RNA molecule with a 5-terminal cap.

An improved data intake and query system that can perform and display ingest-time and search-time field extraction, redaction, copy, and/or categorization is described herein. As described herein, ingest-time field extraction, redaction, copy, and/or categorization may refer to field or field value extraction, redaction, copy, and/or categorization that is performed by a log observer system of the data intake and query system on raw machine data as the raw machine data is ingested or received from a publisher. As described herein, search-time field extraction, redaction, copy, and/or categorization may refer to field or field value extraction, redaction, copy, and/or categorization that is performed by the log observer system and/or other components of the improved data intake and query system on historical raw machine data that has already been ingested and indexed by the improved data intake and query system.

The present invention relates to an immobilized capping enzyme, preferably an immobilized Vaccinia virus capping enzyme. Furthermore, the present invention relates to an immobilized cap-specific nucleoside 2-O-methyltransferase, preferably an immobilized Vaccinia virus cap-specific nucleoside 2-O-methyltransferase. Moreover, the present invention relates to a method for immobilizing said enzymes and to a method of using said enzymes for the addition of a 5-cap structure to RNAs. Moreover, the present invention relates to an enzyme reactor for performing the capping reaction using said immobilized enzymes and the subsequent separation of the 5-capped RNA product. In addition, the present invention relates to a kit comprising the capping enzyme and/or the cap-specific nucleoside 2-O-methyltransferase.

The present disclosure relates to compositions, kits, and methods of making RNA vaccines having an appropriate cap structure. Systems, apparatus, compositions, and/or methods may include and/or use, in some embodiments, non-naturally occurring single-chain RNA capping enzymes. In some embodiments, an RNA capping enzyme may include an FCE variant having (a) an amino acid sequence at least 90% identical to positions 1 to 878 of SEQ ID NO: 1, and/or (b) one or more substitutions relative to SEQ ID NO: 1 at a position selected from positions corresponding to positions 215, 337, 572, 648, and 833 (e.g., a position selected from positions corresponding to position 215, 337, and 572) of SEQ ID NO: 1.

The present disclosure relates to compositions, kits, and methods of making RNA vaccines having an appropriate cap structure. Systems, apparatus, compositions, and/or methods may include and/or use, in some embodiments, non-naturally occurring single-chain RNA capping enzymes. In some embodiments, an RNA capping enzyme may include an FCE variant having (a) an amino acid sequence at least 90% identical to positions 1 to 878 of SEQ ID NO: 1, and/or (b) one or more substitutions relative to SEQ ID NO: 1 at a position selected from positions corresponding to positions 215, 337, 572, 648, and 833 (e.g., a position selected from positions corresponding to position 215, 337, and 572) of SEQ ID NO: 1.

The invention provides a chimeric enzyme comprising at least one catalytic domain of a RNA triphosphatase, at least one catalytic domain of a guanylyltransferase, at least one catalytic domain of a N.sup.7-guanine methyltransferase, and at least one catalytic domain of a DNA-dependant RNA polymerase. The invention also provides pharmaceutical composition comprising said chimeric enzyme and uses of said chimeric enzyme.

Provided herein is a method for efficiently capping RNA in vitro. In some embodiments the capping reaction may be done at high temperature using Vaccinia capping enzyme or a variant thereof. In other embodiments, the capping reactions may comprise a capping enzyme from a large virus of amoeba, e.g., Faustovirus, mimivirus or moumouvirus, or a variant thereof. Compositions and kits for practicing the method are also provided.