Methods for cell free protein synthesis and post translational modification of the expressed proteins
AssigneeNuclera Ltd
InventorsMichael Chun Hao Chen, Gordon Ross Mclnroy
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Abstract
Provided herein are methods of cell-free protein synthesis, optimised cell-free protein synthesis (CFPS) reagents and the post translational modification of the expressed proteins, to increase protein expression yields. The methods are applicable to protein expression on a microfluidic device having hydrophobic surfaces by merging droplets on the device in order to screen a selection of expression and post translational modification compositions in parallel.
Claims21
A method for expressing and changing the molecular weight of an expressed protein using a single reagent composition comprising a template nucleic acid, reagents for cell-free protein synthesis and one or more enzymes which post translationally covalently modify and amend the molecular weight of the expressed protein.
The method according to any one preceding claim, wherein the expression is performed in droplets on an electrowetting-on-dielectric (EWoD) device.
A method for the synthesis of a modified protein on an electrowetting-on-dielectric (EWoD) device, the method comprising taking a reaction system having at least one template nucleic acid encoding a protein of interest, blending droplets to form a series of droplets having varying cell-free reagent compositions, including enzymes for protein synthesis, the template nucleic acid and one or more enzymes for post translationally covalently modifying the expressed folded protein and monitoring the synthesis of the protein of interest in the various compositions, thereby identifying a composition suitable for expression and modification of the protein of interest.
The method according to any one preceding claim, wherein the reagent contains a termination factor.
The method according to any one preceding claim, wherein the reagent composition is formed on an EWoD device by merging a first droplet containing a cell lysate and a second droplet having one or more enzymes for post translationally covalently modifying the expressed folded protein.
The method according to any one preceding claim, wherein the reagent composition is formed on an EWoD device by merging a first droplet containing the template nucleic acid with a second droplet containing the reagent composition having enzymes for protein synthesis and a third droplet having one or more enzymes for post translationally covalently modifying the expressed folded protein.
A composition comprising a template nucleic acid, reagents for cell-free protein synthesis and one or more enzymes for post translationally covalently modifying the expressed folded protein.
Dependencies are inferred from the claim text.