Eukaryotic host system for producing recombinant proteins
Disclosed herein are eukaryotic host systems for producing recombinant proteins, eukaryotic hosts modified to produce recombinant proteins, and methods of producing recombinant proteins using the same. In certain embodiments, the eukaryotic host system is a plasmodium of the class Myxogastria, such as Physarum polycephalum , modified to produce one or more recombinant proteins, such as human insulin. In certain embodiments, a method of protein production includes collecting one or more recombinant proteins produced by Physarum polycephalum while in a form of multinucleate plasmodia.
1 . A system for protein production comprising:
a growth chamber; and
multinucleate plasmodia of Physarum polycephalum disposed within the growth chamber, wherein the Physarum polycephalum is modified to comprise a nucleic acid sequence having at least 90% sequence identity to SEQ ID NO: 1 or SEQ ID NO:2 that, when expressed by the Physarum polycephalum , results in the production of one or more recombinant proteins encoded by the nucleic acid sequence, wherein the one or more recombinant proteins are expressed with a secretion tag, and wherein the growth chamber comprises a port to allow removal of fluid containing the one or more recombinant proteins; and wherein the one or more recombinant proteins are selected from one or more of glucose isomerase, glucose dehydrogenase, dihydroxyacid dehydratase, gluconate/galactonate dehydratase, glyceraldehyde dehydrogenase, pyruvate decarboxylase, alcohol dehydrogenase, functional variants thereof, or combinations thereof.
2 . Physarum polycephalum modified to produce a metabolic enzyme or a functional variant thereof having enzyme activity.
3 . The system of claim 1 , wherein the multinucleate plasmodia are macroplasmodia.
4 . The system of claim 1 , wherein the nucleic acid sequence is chromosomally-integrated into the Physarum polycephalum.
5 . The system of claim 1 , wherein the Physarum polycephalum is diploid when in the form of the multinucleate plasmodia.
6 . The system of claim 1 , wherein the one or more recombinant proteins comprise a metabolic enzyme or a functional variant thereof.
7 . The system of claim 1 , wherein the nucleic acid sequence encodes for glucose isomerase, dihydroxyacid dehydratase, gluconate/galactonate dehydratase, glyceraldehyde dehydrogenase, pyruvate decarboxylase, alcohol dehydrogenase, functional variants thereof, or combinations thereof.
8 . The Physarum polycephalum of claim 2 , in a form of macroplasmodia.
9 . The Physarum polycephalum of claim 2 , comprising a nucleic acid sequence encoding the metabolic enzyme or the functional variant thereof, wherein the nucleic acid sequence is chromosomally-integrated into the Physarum polycephalum.
10 . The Physarum polycephalum of claim 2 , wherein the metabolic enzyme is selected from glucose isomerase, glucose dehydrogenase, dihydroxyacid dehydratase, gluconate/galactonate dehydratase, glyceraldehyde dehydrogenase, pyruvate decarboxylase, alcohol dehydrogenase, functional variants thereof, or combinations thereof.
11 . A system for protein production comprising: a growth chamber; and multinucleate plasmodia of Physarum polycephalum disposed within the growth chamber, wherein the Physarum polycephalum is modified to comprise a nucleic acid sequence having at least 90% sequence identity to SEQ ID NO:1 or SEQ ID NO:2 that, when expressed by the Physarum polycephalum , results in the production of one or more recombinant proteins, wherein the one or more recombinant proteins is selected from glucose isomerase, dihydroxyacid dehydratase, gluconate/galactonate dehydratase, glyceraldehyde dehydrogenase, pyruvate decarboxylase, alcohol dehydrogenase, functional variants thereof, or combinations thereof.
12 . The system of claim 1 , wherein the nucleic acid sequence comprises SEQ ID NO: 1 or SEQ ID NO:2.
13 . The system of claim 1 , wherein the secretion tag is a prespore specific protein A (PsA) signal peptide.
14 . The system of claim 1 , wherein the secretion tag is native to Dictyostelium discoideum.