SEQUENCE LISTING

The instant application contains a Sequence Listing which has been submitted via EFS-web and is hereby incorporated by reference in its entirety. The ASCII copy as filed herewith was originally created on Aug. 27, 2019. The ASCII copy as filed herewith is named NREL 17-47_ST25.txt, is 12 kilobytes in size and is submitted with the instant application.

BACKGROUND

In order to drive innovation in chemical and material applications beyond what has been afforded by the now mature petrochemical industry, new molecules that possess diverse chemical moieties not readily accessed from petroleum are needed. One source of such molecules lies in the varied pathways soil microbes utilize to degrade the abundant aromatic compounds generated during plant decomposition. These pathways may converge at (or “funnel to”) central aromatic intermediates including catechol, protocatechuate, and gallate. The aromatic rings of these compounds may be cleaved by ring-opening dioxygenase enzymes and their products may be metabolized through pathways that eventually enter central carbon metabolism for energy and growth. Intermediate compounds in these pathways provide significant and potentially enabling chemical functionality for biopolymer applications. Thus, there remains a need for improved compositions and methods that more effectively funnel biomass degradation products through pathways to produce intermediate molecules at higher yields, titers, growth rates, and/or efficiencies.

SUMMARY

An aspect of the present disclosure is a non-naturally occurring microorganism that includes an endogenous genetic deletion that eliminates the expression of at least a pyruvate kinase, where the genetically modified prokaryotic microorganism is capable of producing 3-deoxy-D-arabino-heptulosonate-7-phosphate. In some embodiments of the present disclosure, the pyruvate kinase includes at least one of PykA and/or PykF. In some embodiments of the present disclosure, the endogenous genetic deletion may further eliminate the expression of a glucose-6-phosphate isomerase. In some embodiments of the present disclosure, the glucose-6-phosphate isomerase may include at least one of Pgi-1 and/or Pgi-2.

In some embodiments of the present disclosure, the endogenous genetic deletion may further eliminate the expression of a glucose dehydrogenase. In some embodiments of the present disclosure, the glucose dehydrogenase may include Gcd. In some embodiments of the present disclosure, the endogenous genetic deletion may further eliminate the expression of a phosphoenolpyruvate carboxylase. In some embodiments of the present disclosure, the phosphoenolpyruvate carboxylase may include Ppc. In some embodiments of the present disclosure, the endogenous genetic deletion may further eliminate the expression of a transcriptional repressor. In some embodiments of the present disclosure, the transcriptional repressor may include HexR.

In some embodiments of the present disclosure, the genetically modified prokaryotic microorganism may further include an exogenous genetic addition encoding at least one of a DHAP synthase, a decarboxylase, and/or a dehydratase. In some embodiments of the present disclosure, the DHAP synthase may include AroGD146N.

In some embodiments of the present disclosure, the genetically modified prokaryotic microorganism may be from the genusPseudomonas. In some embodiments of the present disclosure, the genetically modified prokaryotic microorganism may be at least one ofP. putida, P. fluorescens, and/orP. stutzeri. In some embodiments of the present disclosure, the genetically modified prokaryotic microorganism may beP. putidaKT2440. In some embodiments of the present disclosure, the genetically modified prokaryotic microorganism may be capable of catabolizing at least one of a lignin depolymerization product, a cellulose depolymerization product, and/or a hemicellulose depolymerization product. In some embodiments of the present disclosure, the lignin depolymerization product may include an aromatic compound. In some embodiments of the present disclosure, the cellulose depolymerization product may include glucose.

In an aspect, disclosed herein is a non-naturally occurring microorganism comprising an endogenous genetic deletion that eliminates the expression of at least a pyruvate kinase, wherein the microorganism is capable of producing 3-deoxy-D-arabino-heptulosonate-7-phosphate. In an embodiment, the microorganism has a pyruvate kinase that comprises at least one of PykA or PykF. In an embodiment, the microorganism has an endogenous genetic deletion that further eliminates the expression of a glucose-6-phosphate isomerase. In an embodiment, the microorganism has glucose-6-phosphate isomerase that comprises at least one of Pgi-1 or Pgi-2. In another embodiment, the microorganism contains an endogenous genetic deletion that further eliminates the expression of a glucose dehydrogenase. In an embodiment, the microorganism has glucose dehydrogenase that comprises Gcd. In an embodiment, the microorganism has an endogenous genetic deletion that eliminates the expression of a phosphoenolpyruvate carboxylase. In another embodiment, the microorganism has a phosphoenolpyruvate carboxylase that comprises Ppc. In an embodiment, the microorganism contains an endogenous genetic deletion that further eliminates the expression of a transcriptional repressor. In another embodiment, the microorganism has a deletion of the transcriptional repressor comprises HexR. In an embodiment, the microorganism has an exogenous genetic addition encoding at least one of a DHAP synthase, a decarboxylase, or a dehydratase. In another embodiment, the microorganism has a DHAP synthase that comprises AroGD146N. In yet another embodiment, the microorganism of is a genetically modified prokaryotic microorganism from the genusPseudomonas. In yet another embodiment, the microorganism is selected from the group consisting ofP. putida, P. fluorescens, andP. stutzeri. In an embodiment, the microorganism isP. putidaKT2440. In yet another embodiment, the microorganism is capable of catabolizing at least one of a lignin depolymerization product, a cellulose depolymerization product, or a hemicellulose depolymerization product. In another embodiment, the lignin depolymerization product comprises an aromatic compound. In an embodiment, the cellulose depolymerization product comprises glucose. In another embodiment, the microorganism has a deletion of gacS. In an embodiment, the microorganism of has a deletion of gnd.

