	US 12,460,234 B2
	Synthesis of isoprenoids and derivatives
Ramon Gonzalez, Tampa, FL (US); James M. Clomburg, Houston, TX (US); and Seokjung Cheong, Emeryville, CA (US)
Assigned to WILLIAM MARSH RICE UNIVERSITY, Houston, TX (US)
Filed by William Marsh Rice University, Houston, TX (US)
Filed on Jun. 7, 2023, as Appl. No. 18/330,944.
Application 18/330,944 is a division of application No. 17/234,398, filed on Apr. 19, 2021, abandoned.
Application 17/234,398 is a division of application No. 16/081,756, granted, now 11,046,978, issued on Jun. 29, 2021, previously published as PCT/US2017/022581, filed on Mar. 15, 2017.
Claims priority of provisional application 62/308,937, filed on Mar. 16, 2016.
Claims priority of provisional application 62/343,598, filed on May 31, 2016.
Prior Publication US 2024/0182932 A1, Jun. 6, 2024
Int. Cl. C12P 7/42 (2006.01)

CPC C12P 7/42 (2013.01)

8 Claims

1. A recombinant bacteria for synthesizing an isoprenoid, the recombinant bacteria comprising the following overexpressed enzymes:

a) a thiolase capable of condensing two acetyl-CoAs to form acetoacetyl CoA;

b) an overexpressed 3-hydroxy-3-methylglutaryl-CoA synthase capable of condensing said acetoacetyl-CoA with acetyl-CoA to form 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA);

c) an enoyl-CoA hydratase capable of dehydrating said HMG-CoA to 3-methylglutaconylCoA;

d) a glutaconyl-CoA decarboxylase capable of decarboxylating said 3-methylglutaconyl-CoA to 3-methylcrotonyl-CoA (aka 3-methyl-2-butenoyl-CoA);

e) one or more enzyme(s) catalyzing conversion of said 3-methylcrotonyl-CoA to prenol (aka 3-methyl, 2-butenol), wherein said enzyme(s) are selected from:

i) an alcohol-forming acyl-CoA reductase;

ii) an aldehyde forming acyl-CoA reductase plus an alcohol dehydrogenase; or

iii) a carboxylate reductase plus an alcohol dehydrogenase plus an enzyme(s) selected from an acyl-CoA synthase, an acyl-CoA transferase, a thioesterase, or a carboxylate kinase plus a phosphotransacylase;

f) one or more enzyme(s) to convert said prenol to dimethylallyl pyrophosphate (DMAPP) and isopentenyl pyrophosphate (IPP), said enzyme(s) selected from:

i) an alcohol kinase (EC 2.7.1.-) plus a phosphate kinase (EC 2.7.4.-); or

ii) an alcohol diphosphokinase (EC 2.7.6.-) plus an IPP isomerase (5.3.3.2);

g) a GPP synthase capable of catalyzing condensation of said DMAPP and IPP to form geranyl diphosphate (GPP); and

h) a terpene synthase (EC 4.2.3.-) capable of converting said GPP to an isoprenoid.