Compositions and methods for rapid and dynamic flux control using synthetic metabolic valves

The genetically modified microorganism claim 1, wherein the genetically modified microorganism is capable of expression a heterologous enzyme of a product production pathway and the expression of the heterologous enzyme is conditional and/or coordinated with the conditional triggering of the SMV.

The genetically modified microorganism claim 1, wherein the heterologous enzyme is selected from the group consisting of:

• an acetoacetyl-CoA thiolase, hydroxymethylglutaryl-CoA synthase, hydroxymethylglutaryl-CoA reductase, mevalonate kinase, phosphomevalonate kinase, mevalonte diphosphate decarboxylase, isopentenyl-diphosphate isomerase, isoprene synthase and combinations thereof.

The genetically modified microorganism claim 1, wherein the heterologous enzymes are mvaE gene from Enterococcus faecalis and the mvaS gene, also from E. faecalis.

The genetically modified microorganism claim 1, wherein the production pathway comprises both increased expression of an acetyl-CoA carboxylase enzyme and increased expression of an enzyme of an isoprenoid production pathway.

The genetically modified microorganism of claim 1, wherein the first and second enzyme essential for growth of the genetically modified microorganism are each selected from the group consisting of:

• enoyl-ACP reductase (fabI), citrate synthase (gltA), soluble transhydrogenase (udhA), glucose-6-phosphate-1-dehydrogenase (zwf), lipoamide dehydrogenase (lpd), and combinations thereof.

The genetically modified microorganism of claim 1, wherein the first and second enzyme essential for growth of the genetically modified microorganism are each selected from the group consisting of:

• phosphoenolpyruvate carboxylase (ppc), succinyl-CoA synthetase (sucD), isocitrate lyase (aceA), ATP-dependent 6-phosphofructokinase (pfkA), ATP-dependent Lon proteinse (lon), sigma factor sigma-38 (rpoS), transketolase 1 (tktA), transketolast 2 (tktB), and combinations thereof.

The genetically modified microorganism of claim 1, wherein the synthetic metabolic valve comprises:

• (i) a gene encoding more than one small guide RNA specific for targeting more than one gene of an enzyme essential for growth of the genetically modified microorganism;
• (ii) a gene encoding a Cas9 or a gene encoding a dCAS9, and
• (iii) a gene encoding a proteolytic enzyme enhancing factor sspB.

The genetically modified microorganism claim 1, wherein the limiting nutrient of the growth media is a nutrient selected from the group consisting of phosphate, nitrogen, sulfur, and magnesium, or any combinations thereof.

The genetically modified microorganism claim 1, wherein the genetically modified microorganism may be at least partially controlled by chemical inducer that is selected from a group consisting of tetracycline, anhydrotetracycline, lactose, IPTG (isopropyl-beta-D-1-thiogalactopyranoside), arabinose, raffinose, and tryptophan, and combinations thereof.

The genetically modified microorganism of claim 1, wherein the genetically modified microorganism further comprises a disruption or deletion of a gene naturally occurring in the genetically modified mircroorganism and the gene selected from the group consisting of a gene encoding lactate dehydrogenase (ldhA), phosphate acetyltransferase (pta), pyruvate oxidase (poxB), pyruvate-formate lyase (pflB), the methylglyoxal synthase (mgsA), acetate kinase (ackA), alcohol dehydrogenase (adhE), ATP-dependent Lon protease (lon), outer membrane protease (ompT), arcA transcriptional dual regulator (arcA), iclR transcriptional regulator (iclR), and any combinations thereof.