41 research outputs found
Recommended from our members
Module-Based Polyketide Synthase Engineering for <i>de Novo</i> Polyketide Biosynthesis
Polyketide retrobiosynthesis, where the biosynthetic pathway of a given polyketide can be reversibly engineered due to the colinearity of the polyketide synthase (PKS) structure and function, has the potential to produce millions of organic molecules. Mixing and matching modules from natural PKSs is one of the routes to produce many of these molecules. Evolutionary analysis of PKSs suggests that traditionally used module boundaries may not lead to the most productive hybrid PKSs and that new boundaries around and within the ketosynthase domain may be more active when constructing hybrid PKSs. As this is still a nascent area of research, the generality of these design principles based on existing engineering efforts remains inconclusive. Recent advances in structural modeling and synthetic biology present an opportunity to accelerate PKS engineering by re-evaluating insights gained from previous engineering efforts with cutting edge tools
Recommended from our members
Leveraging microbial biosynthetic pathways for the generation of 'drop-in' biofuels.
Advances in retooling microorganisms have enabled bioproduction of 'drop-in' biofuels, fuels that are compatible with existing spark-ignition, compression-ignition, and gas-turbine engines. As the majority of petroleum consumption in the United States consists of gasoline (47%), diesel fuel and heating oil (21%), and jet fuel (8%), 'drop-in' biofuels that replace these petrochemical sources are particularly attractive. In this review, we discuss the application of aldehyde decarbonylases to produce gasoline substitutes from fatty acid products, a recently crystallized reductase that could hydrogenate jet fuel precursors from terpene synthases, and the exquisite control of polyketide synthases to produce biofuels with desired physical properties (e.g., lower freezing points). With our increased understanding of biosynthetic logic of metabolic pathways, we discuss the unique advantages of fatty acid, terpene, and polyketide synthases for the production of bio-based gasoline, diesel and jet fuel
Sulfonyl 3鈥慉lkynyl Pantetheinamides as Mechanism-Based Cross-Linkers of Acyl Carrier Protein Dehydratase
Acyl carrier proteins (ACPs) play
a central role in acetate biosynthetic
pathways, serving as tethers for substrates and growing intermediates.
Activity and structural studies have highlighted the complexities
of this role, and the protein鈥損rotein interactions of ACPs
have recently come under scrutiny as a regulator of catalysis. As
existing methods to interrogate these interactions have fallen short,
we have sought to develop new tools to aid their study. Here we describe
the design, synthesis, and application of pantetheinamides that can
cross-link ACPs with catalytic 尾-hydroxy-ACP dehydratase (DH)
domains by means of a 3-alkynyl sulfone warhead. We demonstrate this
process by application to the Escherichia coli fatty acid synthase and apply it to probe protein鈥損rotein
interactions with noncognate carrier proteins. Finally, we use solution-phase
protein NMR spectroscopy to demonstrate that sulfonyl 3-alkynyl pantetheinamide
is fully sequestered by the ACP, indicating that the <i>crypto</i>-ACP closely mimics the natural DH substrate. This cross-linking
technology offers immediate potential to lock these biosynthetic enzymes
in their native binding states by providing access to mechanistically
cross-linked enzyme complexes, presenting a solution to ongoing structural
challenges
Probing the selectivity and protein路protein interactions of a nonreducing fungal polyketide synthase using mechanism-based crosslinkers.
Protein路protein interactions, which often involve interactions among an acyl carrier protein (ACP) and ACP partner enzymes, are important for coordinating polyketide biosynthesis. However, the nature of such interactions is not well understood, especially in the fungal nonreducing polyketide synthases (NR-PKSs) that biosynthesize toxic and pharmaceutically important polyketides. Here, we employ mechanism-based crosslinkers to successfully probe ACP and ketosynthase (KS) domain interactions in NR-PKSs. We found that crosslinking efficiency is closely correlated with the strength of ACP路KS interactions and that KS demonstrates strong starter unit selectivity. We further identified positively charged surface residues by KS mutagenesis, which mediates key interactions with the negatively charged ACP surface. Such complementary/matching contact pairs can serve as "adapter surfaces" for future efforts to generate new polyketides using NR-PKSs
Recommended from our members
Production of Odd-Carbon Dicarboxylic Acids in Escherichia coli Using an Engineered Biotin-Fatty Acid Biosynthetic Pathway.
Dicarboxylic acids are commodity chemicals used in the production of plastics, polyesters, nylons, fragrances, and medications. Bio-based routes to dicarboxylic acids are gaining attention due to environmental concerns about petroleum-based production of these compounds. Some industrial applications require dicarboxylic acids with specific carbon chain lengths, including odd-carbon species. Biosynthetic pathways involving cytochrome P450-catalyzed oxidation of fatty acids in yeast and bacteria have been reported, but these systems produce almost exclusively even-carbon species. Here we report a novel pathway to odd-carbon dicarboxylic acids directly from glucose in Escherichia coli by employing an engineered pathway combining enzymes from biotin and fatty acid synthesis. Optimization of the pathway will lead to industrial strains for the production of valuable odd-carbon diacids
Production of Odd-Carbon Dicarboxylic Acids in <i>Escherichia coli</i> Using an Engineered Biotin鈥揊atty Acid Biosynthetic Pathway
Dicarboxylic acids are commodity
chemicals used in the production
of plastics, polyesters, nylons, fragrances, and medications. Bio-based
routes to dicarboxylic acids are gaining attention due to environmental
concerns about petroleum-based production of these compounds. Some
industrial applications require dicarboxylic acids with specific carbon
chain lengths, including odd-carbon species. Biosynthetic pathways
involving cytochrome P450-catalyzed oxidation of fatty acids in yeast
and bacteria have been reported, but these systems produce almost
exclusively even-carbon species. Here we report a novel pathway to
odd-carbon dicarboxylic acids directly from glucose in <i>Escherichia
coli</i> by employing an engineered pathway combining enzymes
from biotin and fatty acid synthesis. Optimization of the pathway
will lead to industrial strains for the production of valuable odd-carbon
diacids