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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 73 records · Page 4
All sourcesNASA NTRSDOE OSTI
DOE OSTI2018

Dietary nutrient levels alter the metabolism of arginine family amino acids in the conceptus of Huanjiang mini‐pigs

Abstract BACKGROUND The arginine family amino acids (AFAAs) exert important roles in the metabolism, growth and development of the conceptus. However, to date, few studies have investigated the effects of maternal nutrient levels on the concentrations and metabolism of AFAAs in the conceptus. RESULTS Compared to low nutrient diets, high nutrient diets increased ( P < 0.05) the concentrations of citrulline and proline (Pro) in plasma; the concentrations of arginine, glutamine, Pro and ornithine (Orn) in the amniotic fluid; and the concentrations of all detected AFAAs in the allantoic fluid, which were most pronounced on day 45 of pregnancy. High nutrient diets upregulated ( P < 0.05) mRNA expression of arginase I ( Arg I ), Pro oxidase and spermidine synthetase ( SRM ) in the fetal placenta, as well as Arg II , SRM and spermine synthetase ( SMS ) expression in the fetal liver (most pronounced on day 45 of pregnancy). The same effect was observed for mRNA expression of NO synthase and Orn aminotransferase ( OAT ), mainly on day 110 of pregnancy, and for mRNA expression of Arg I , Arg II , OAT , Orn decarboxylase and SMS throughout pregnancy. High nutrient diets upregulated ( P < 0.05) mRNA expression of Y + L‐type amino acid transporter ( LAT ) and cationic amino acid transporter 1 ( CAT1 ) in the fetal jejunum throughout pregnancy. Dietary treatments did not affect ( P > 0.05) mRNA expression of Y + LAT1 , sodium‐coupled neutral amino acid transporter 2 ( SNAT2 ) and CAT1 in the fetal placenta, skeletal muscle and colon. CONCLUSION High nutrient diets increased the concentration and transport of AFAAs in the mothers and conceptus, which likely improves growth and development of the conceptus. © 2018 Society of Chemical Industry

Duan, Yehui↗
DOE OSTI2020

Regulatory interventions improve the biosynthesis of limiting amino acids from methanol carbon to improve synthetic methylotrophy in Escherichia coli

Synthetic methylotrophy aims to engineer methane and methanol utilization pathways in platform hosts like Escherichia coli for industrial bioprocessing of natural gas and biogas. While recent attempts to engineer synthetic methylotrophs have proved successful, autonomous methylotrophy, that is, the ability to utilize methane or methanol as sole carbon and energy substrates, has not yet been realized. Here, we address an important limitation of autonomous methylotrophy in E. coli: the inability of the organism to synthesize several amino acids when grown on methanol. We targeted global and local amino acid regulatory networks. Those include removal of amino acid allosteric feedback inhibition (argA H15Y , ilvA L447F , hisG E271K , leuA G462D , proB D107N , thrA S345F, trpE S40F ), knockouts of transcriptional repressors (ihfA, metJ); and overexpression of amino acid biosynthetic operons (hisGDCBHAFI, leuABCD, thrABC, trpEDCBA) and transcriptional regulators (crp, purR). Compared to the parent methylotrophic E. coli strain that was unable to synthesize these amino acids from methanol carbon, these strategies resulted in improved biosynthesis of limiting proteinogenic amino acids (histidine, leucine, lysine, methionine, phenylalanine, threonine, tyrosine) from methanol carbon. In several cases, improved amino acid biosynthesis from methanol carbon led to improvements in methylotrophic growth in methanol minimal medium supplemented with a small amount of yeast extract. Here, this study addresses a key limitation currently preventing autonomous methylotrophy in E. coli and possibly other synthetic methylotrophs and provides insight as to how this limitation can be alleviated via global and local regulatory modifications.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Understanding the Nature of Amino Acid Interactions with Pd(111) or Pd–Au Bimetallic Catalysts in the Aqueous Phase

In this study, the interaction of methionine (Met) with different bimetallic-segregated surfaces comprising a uniform distribution of strips and islands of Au on the Pd(111) surface was examined using molecular dynamics (MD) simulations. Out of all the segregated and uniformly doped surfaces studied, the design of Pd–Au islands showed some reduction in the interaction energy (E int = -43.7 kJ/mol) as compared to that of the pure Pd(111) surface (E int = -50 kJ/mol) for a single Met molecule. However, at a higher coverage of 9 Met molecules/simulation cell, none of the Pd–Au alloy surfaces showed any improvement as compared to the Pd(111) surface. In order to develop a comprehensive understanding of the nature of the nonbonded interaction of aqueous biogenic impurities with the Pd catalyst surface, the MD study was extended to include a variety of aliphatic, S-containing, aromatic, and polar amino acids. The potential of mean force (PMF) profiles were observed to be distinct for each class of amino acids with substantial differences among amino acids with acidic and basic side chains. The side chains of all the polar and aromatic amino acids showed direct contact with the surface while aliphatic amino acids had their hydrophobic side chain aligned away from the surface. Interestingly, lysine (Lys) and tyrosine (Tyr) were the only two amino acids which interacted preferentially via the distant backbone nitrogen and backbone oxygen, respectively, despite their side chains being in direct contact with the metal surface. The strength of interaction was correlated with the size of the amino acid; the interaction energies were observed to be the maximum for large molecules such as arginine (Arg, E int = -87.7 kJ/mol) and tryptophan (Trp, E int = -73.4 kJ/mol), while it was a minimum for aliphatic amino acids such as alanine (Ala, E int = -10.9 kJ/mol). Lastly, the study is focused on examining the sensitivity of the choice of the preferential interaction site, conformational preferences, and interaction energies to the side-chain specificity.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2021

Amino Acids Dominate Diffusive Nitrogen Fluxes in Acidic Tussock Tundra

Organic nitrogen (N) is abundant in soils, but early conceptual frameworks considered it nonessential for plant growth. It is now well recognised that plants have the potential to take up organic N. However, it is still unclear whether plants supplement their N requirements by taking up organic N in situ: at what rate is organic N diffusing towards roots and are plants taking it up? In this work, we combined microdialysis with live-root uptake experiments to measure amino acid speciation and diffusion rates towards roots of Eriophorum vaginatum. Amino acid diffusion rates (321 ng N cm -2 h -1 ) were c. 3× higher than those for inorganic N. Positively charged amino acids made up 68% of the N diffusing through soils compared with neutral and negatively charged amino acids. Live-root uptake experiments confirmed that amino acids are taken up by plants (up to 1 µg N g -1 min -1 potential net uptake). Amino acids must be considered when forecasting plant-available N, especially when they dominate the N supply, and when acidity favours proteolysis over net N mineralisation. Determining amino acid production pathways and supply rates will become increasingly important in projecting the extent and consequences of shrub expansion, especially considering the higher C : N ratio of plants relative to soil.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2019

Real‐time quantification and supplementation of bioreactor amino acids to prolong culture time and maintain antibody product quality

Abstract Real‐time monitoring of cell cultures in bioreactors can enable expedited responses necessary to correct potential batch failure perturbations which may normally go undiscovered until the completion of the batch and result in failure. Currently, analytical technologies are dedicated to real‐time monitoring of bioreactor parameters such as pH, dissolved oxygen, and temperature, nutrients such as glucose and glutamine, or metabolites such as lactate. Despite the importance of amino acids as the building blocks of therapeutic protein products, other than glutamine their concentrations are not commonly measured. Here, we present a study into amino acid monitoring, supplementation strategies, and how these techniques may impact the cell growth profiles and product quality. We used preliminary bioreactor runs to establish baselines by determining initial amino acid consumption patterns, the results of which were used to select a pool of amino acids which gets depleted in the bioreactor. These amino acids were combined into blends which were supplemented into bioreactors during a subsequent run, the concentrations of which were monitored using a mass spectrometry based at‐line method we developed to quickly assess amino acid concentrations from crude bioreactor media. We found that these blends could prolong culture life, reversing a viable cell density decrease that was leading to batch death. Additionally, we assessed how these strategies might impact protein product quality, such as the glycan profile. The amino acid consumption data were aligned with the final glycan profiles in principal component analysis to identify which amino acids are most closely associated with glycan outcomes.

Powers, David N.↗
DOE OSTI2023

Examining Interactions of Uranyl(VI) Ions with Amino Acids in the Gas Phase

Gas-phase experiments, using electrospray ionization quadrupole ion trap mass spectrometry (ESI-QIT/MS), were conducted to probe basic interactions of the uranyl(VI) ion, UO 2 2+ , with selected natural amino acids, namely, L-cysteine (Cys), L-histidine (His), and L-aspartic acid (Asp), which strongly bind to metal ions. The simplest amino acid, glycine (Gly), was also studied for comparison. Cys, His, and Asp have additional potentially coordinating groups beyond the amino and carboxylic acid functional groups, specifically thiol in Cys, imidazole in His, and a second carboxylate in Asp. Gas-phase experiments comprised collision-induced dissociation (CID) of uranyl–amino acid complexes and competitive CID to assess the relative binding strength of different amino acids in the same uranyl complex. Reactivity of selected uranyl–amino acid complexes with water provided further insights into relative stabilities. In positive ion mode, CID and ensuing reactions with water suggested that uranyl–neutral AA binding strength decreased in the order His > Asp > Cys > Gly, which is similar to amino acid proton affinities. In negative ion mode, CID revealed a decreasing dissociation tendency in the order Gly >> His ≈ Cys > Asp, presumably reflecting a reverse enhanced binding to uranyl of the doubly deprotonated amino acids formed in CID.

38 RADIATION CHEMISTRY, RADIOCHEMISTRY, AND NUCLEA↗
DOE OSTI2017

Biopolymers Containing Unnatural Amino Acids

Although the main chain structure of polymers has a profound effect on their materials properties, the side groups can also have dramatic effects on their properties including conductivity, liquid crystallinity, hydrophobicity, elasticity and biodegradability. Unfortunately control over the side chain structure of polymers remains a challenge – it is difficult to control the sequence of chain elongation when mixtures of monomers are polymerized, and postpolymerization side chain modification is made difficult by polymer effects on side chain reactivity. In contrast, the mRNA templated synthesis of polypeptides on the ribosome affords absolute control over the primary sequence of the twenty amino acid monomers. Moreover, the length of the biopolymer is precisely controlled as are sites of crosslinking. However, whereas synthetic polymers can be synthesized from monomers with a wide range of chemically defined structures, ribosomal biosynthesis is largely limited to the 20 canonical amino acids. For many applications in material sciences, additional building blocks would be desirable, for example, amino acids containing metallocene, photoactive, and halogenated side chains. To overcome this natural constraint we have developed a method that allows unnatural amino acids, beyond the common twenty, to be genetically encoded in response to nonsense or frameshift codons in bacteria, yeast and mammalian cells with high fidelity and good yields. Here we have developed methods that allow identical or distinct noncanonical amino acids to be incorporated at multiple sites in a polypeptide chain, potentially leading to a new class of templated biopolymers. We have also developed improved methods for genetically encoding unnatural amino acids. In addition, we have genetically encoded new amino acids with novel physical and chemical properties that allow selective modification of proteins with synthetic agents. Finally, we have evolved new metal-ion binding sites in proteins using a novel metal-ion binding amino acid, which may facilitate our ability to generate new protein based sensors and catalysts.

36 MATERIALS SCIENCE↗
DOE OSTI2015

Tuning the formations of metal-1,3,5-benzenetricarboxylate frameworks via the assistance of amino acids

Seven new metal-1,3,5-benzenetricarboxylate coordination polymers have been synthesized by modification of auxiliary components during the assembly reactions. Their structures have been determined by single-crystal X-ray diffraction analyses and further characterized by XRD and TGA. Interestingly, they show fascinating topological structures. Compounds 1 and 2 possess the undulating layer structure with 3-connected hcb network and (3,6)-connected kgd network. Compound 3 possesses three-dimensional (3D) pillared-layer structure with 3-connected 2-fold interpenetrating srs net. Compound 4 also has the 3D 2-fold interpenetrating pillared-layer structure; however, it has (3,5)-connected hms topology because the Cd(II) center is 5-connected. Compound 5 possess 3D structure through hydrogen bonding interactions between ladder-like layers. Compounds 6 and 7 have the similar 3D frameworks with 4-connected umc net and (3,7)-connected (3.4.5)(3{sup 2}.4{sup 6}.5{sup 5}.6{sup 8}) topology, respectively. The photoluminescent properties of compounds 2–7 were also investigated. - Graphical abstract: Presented here are seven new metal-1,3,5-benzenetricarboxylate coordination polymers with diverse structures from 2D layers to 3D open frameworks. The synthesis and structural diversity of these compounds are determined by the additional amino acids as unusual buffering agents. - Highlights: • Structural diversity of metal-1,3,5-benzenetricarboxylate frameworks. • Tuning structural topologies of MOFs via the assistance of amino acids. • Amino acids as unusual buffering agents for the synthesis of MOFs.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2014

Enantiomeric excesses induced in amino acids by ultraviolet circularly polarized light irradiation of extraterrestrial ice analogs: A possible source of asymmetry for prebiotic chemistry

The discovery of meteoritic amino acids with enantiomeric excesses of the L-form (ee {sub L}) has suggested that extraterrestrial organic materials may have contributed to prebiotic chemistry and directed the initial occurrence of the ee {sub L} that further led to homochirality of amino acids on Earth. A proposed mechanism for the origin of ee {sub L} in meteorites involves an asymmetric photochemistry of extraterrestrial ices by UV circularly polarized light (CPL). We have performed the asymmetric synthesis of amino acids on achiral extraterrestrial ice analogs by VUV CPL, investigating the chiral asymmetry transfer at two different evolutionary stages at which the analogs were irradiated (regular ices and/or organic residues) and at two different photon energies (6.6 and 10.2 eV). We identify 16 distinct amino acids and precisely measure the L-enantiomeric excesses using the enantioselective GC × GC-TOFMS technique in five of them: α-alanine, 2,3-diaminopropionic acid, 2-aminobutyric acid, valine, and norvaline, with values ranging from ee {sub L} = –0.20% ± 0.14% to ee {sub L} = –2.54% ± 0.28%. The sign of the induced ee {sub L} depends on the helicity and the energy of CPL, but not on the evolutionary stage of the samples, and is the same for all five considered amino acids. Our results support an astrophysical scenario in which the solar system was formed in a high-mass star-forming region where icy grains were irradiated during the protoplanetary phase by an external source of CPL of a given helicity and a dominant energy, inducing a stereo-specific photochemistry.

79 ASTRONOMY AND ASTROPHYSICS↗
DOE OSTI2015

Structure of the ordered hydration of amino acids in proteins: analysis of crystal structures

The hydration of protein crystal structures was studied at the level of individual amino acids. The dependence of the number of water molecules and their preferred spatial localization on various parameters, such as solvent accessibility, secondary structure and side-chain conformation, was determined. Crystallography provides unique information about the arrangement of water molecules near protein surfaces. Using a nonredundant set of 2818 protein crystal structures with a resolution of better than 1.8 Å, the extent and structure of the hydration shell of all 20 standard amino-acid residues were analyzed as function of the residue conformation, secondary structure and solvent accessibility. The results show how hydration depends on the amino-acid conformation and the environment in which it occurs. After conformational clustering of individual residues, the density distribution of water molecules was compiled and the preferred hydration sites were determined as maxima in the pseudo-electron-density representation of water distributions. Many hydration sites interact with both main-chain and side-chain amino-acid atoms, and several occurrences of hydration sites with less canonical contacts, such as carbon–donor hydrogen bonds, OH–π interactions and off-plane interactions with aromatic heteroatoms, are also reported. Information about the location and relative importance of the empirically determined preferred hydration sites in proteins has applications in improving the current methods of hydration-site prediction in molecular replacement, ab initio protein structure prediction and the set-up of molecular-dynamics simulations.

75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND↗
DOE OSTI2021

A Process Intensification Approach for CO 2 Absorption Using Amino Acid Solutions and a Guanidine Compound

Environmentally friendly amino-acid salt solutions are used for the absorption of carbon dioxide from concentrated flue-gas streams via chemical absorption. Process intensification reduces operating and capital costs by combining chemical reactions and separation operations. Here, we present a new process-intensification approach that combines the CO 2 capture and the amino-acid regeneration steps into a single process carried out in a slurry three-phase reactor. The absorbed CO 2 precipitates as a solid carbonated guanidine compound. The cycle is completed by separation of the solid precipitate to strip the CO 2 and regenerate the guanidine compound, while the liquid solution is recycled to the slurry reactor. The process was studied by modifying a model developed by the authors for a gas-liquid bubble column without the presence of the guanidine compound. The guanidine precipitation reaction was accounted for using kinetic parameters calculated by the authors in another study. The proposed model was implemented by modifying an existing computer code used for the simulation of gas-liquid bubble columns. The calculated results showed that the proposed cycle can significantly reduce energy, equipment, and operating costs and can make an important contribution to developing a competitive cost-effective large-scale process for CO 2 capture.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2020

Structural basis for divergent and convergent evolution of catalytic machineries in plant aromatic amino acid decarboxylase proteins

Radiation of the plant pyridoxal 5'-phosphate (PLP)-dependent aromatic L -amino acid decarboxylase (AAAD) family has yielded an array of paralogous enzymes exhibiting divergent substrate preferences and catalytic mechanisms. Plant AAADs catalyze either the decarboxylation or decarboxylation-dependent oxidative deamination of aromatic L -amino acids to produce aromatic monoamines or aromatic acetaldehydes, respectively. These compounds serve as key precursors for the biosynthesis of several important classes of plant natural products, including indole alkaloids, benzylisoquinoline alkaloids, hydroxycinnamic acid amides, phenylacetaldehyde-derived floral volatiles, and tyrosol derivatives. In this paper, we present the crystal structures of four functionally distinct plant AAAD paralogs. Through structural and functional analyses, we identify variable structural features of the substrate-binding pocket that underlie the divergent evolution of substrate selectivity toward indole, phenyl, or hydroxyphenyl amino acids in plant AAADs. Moreover, we describe two mechanistic classes of independently arising mutations in AAAD paralogs leading to the convergent evolution of the derived aldehyde synthase activity. Applying knowledge learned from this study, we successfully engineered a shortened benzylisoquinoline alkaloid pathway to produce (S)-norcoclaurine in yeast. This work highlights the pliability of the AAAD fold that allows change of substrate selectivity and access to alternative catalytic mechanisms with only a few mutations.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Chemical design of pH-sensitive nanovalves on the outer surface of mesoporous silicas for controlled storage and release of aromatic amino acid

Mesoporous silicas with hexagonally arranged pore channels were synthesized in water–ethanol-ammonia solution using cetyltrimethylammonium bromide as template. Directed modification of silica surface with N-[N′-(N′-phenyl)-2-aminophenyl]-3-aminopropyl groups was realized by postsynthetic activation of halogenoalkylsilicas, which have surface uniformly or selectively distributed 3-chloropropyl groups, with 2-aminodiphenylamine in the liquid phase. Chemical composition of silica materials was estimated by IR spectroscopy and chemical analysis of the surface products of reactions. Characteristics of porous structure of MCM-41-type silicas were determined from X-ray and low-temperature nitrogen ad-desorption measurements. Release ability of synthesized silica carriers was established on encapsulation of 4-aminobenzoic acid in pore channels and subsequent delivery at pH=6.86 and pH=1.00. It was found that N-[N′-(N′-phenyl)-2-aminophenyl]-3-aminopropyl groups block pore entrances at neutral pH preventing 4-aminobenzoic acid release. At pH=1.00 repulsion of positively charged surface aromatic amino groups localized near pore orifices provides unhindered liberation of aromatic amino acid from mesoporous channels. - Graphical abstract: Blocking of pores with N-[N′-(N′-phenyl)-2-aminophenyl]-3-aminopropyl groups at pH=6.86 for storage of ABA and opening of pore entrances at pH=1.00 for unhindered ABA liberation. - Highlights: • Modification of MCM-41 with N-[N′-(N′-phenyl)-2-aminophenyl]-3-aminopropyl groups. • Study of release ability of synthesized silica carriers in relation to amino acid. • Controlled blocking and opening of pores by amino groups at pH change were performed. • Retention of amino acid at pH=6.86 and its liberation at pH=1.00 was proved.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2016

Structural Biology of Aromatic Amino Acid Biosynthesis in Plants: The Prephenate Branch Point

Plants are chemists that have spent the last 425 million years evolving unique secondary metabolites to communicate with their surroundings, defend themselves against pathogens, insects, and herbivores, and grow and develop. Many secondary metabolites are biosynthesized from the aromatic amino acids tryptophan, tyrosine, and phenylalanine, and the shikimate and aromatic amino acid biosynthetic pathways link primary and specialized metabolism. Prephenate aminotransferase (PAT) is a key enzyme in aromatic amino acid biosynthesis because its product, arogenate, is the immediate precursor for both tyrosine and phenylalanine. Interestingly, the enzyme has a dual role in metabolism and can also function as a classical aspartate aminotransferase. AtPAT is pyridoxal 5′‐phosphate‐dependent (PLP) enzyme belonging to the AAT‐like aspartate aminotransferase domain and follows a ping‐pong bi‐bi reaction mechanism through Schiff‐base formation to a catalytic K306 in the active site. Each subunit of the AtPAT dimer consists of a large domain and a small domain that belong to the α‐β class of protein fold. Despite the structural similarities to aspartate aminotransferases, the molecular basis of prephenate binding specificity remains elusive. To gain an understanding of the enzyme's chemistry and substrate selectivity, a 2.5 Å x‐ray structure of PAT from the model plant Arabidopsis thaliana (thale cress) was solved in complex with the PLP. To our knowledge, this is the first structure of a bona fide prephenate aminotransferase. Amino acid sequence comparisons of all known functional PATs shows that T84 and K169 are highly conserved; however, some enzymes annotated as PAT display sequence variations at these positions. Biochemical analysis of T84V, K169V, and T84V/K169V PAT reveals that the double mutant gained tyrosine aminotransferase activity and was able to convert 4‐hydroxyphenylpyruvate to tyrosine. T84 is on the opposite side of the active site pocket, while the amine backbone of K169 hydrogen bonds to PLP. We hypothesize that Thr84 and Lys169 are important for substrate recognition in the active site and will use these mutants in future crystallographic experiments. Overall, our structural studies will assist in understanding the role of this bi‐functional enzyme in the context of aromatic amino acid biosynthesis in plants. Support or Funding Information This material is based upon work supported by the National Science Foundation Graduate Research Fellowship under Grant No. DGE‐1143954. Any opinion, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the National Science Foundation. Results shown in this report are derived from work performed at Argonne National Laboratory, Structural Biology Center at the Advanced Photon Source. Argonne is operated by UChicago Argonne, LLC, for the U.S. Department of Energy, Office of Biological and Environmental Research under contract DE‐AC02‐06CH11357.

Holland, Cynthia K.↗
DOE OSTI2019

Methyl ketone production by Pseudomonas putida is enhanced by plant-derived amino acids

Plants are an attractive source of renewable carbon for conversion to biofuels and bio-based chemicals. Conversion strategies often use a fraction of the biomass, focusing on sugars from cellulose and hemicellulose. Strategies that use plant components, such as aromatics and amino acids, may improve the efficiency of biomass conversion. Pseudomonas putida is a promising host for its ability to metabolize a wide variety of organic compounds. P. putida was engineered to produce methyl ketones, which are promising diesel blendstocks and potential platform chemicals, from glucose and lignin-related aromatics. Unexpectedly, P. putida methyl ketone production using Arabidopsis thaliana hydrolysates was enhanced 2-5-fold compared with sugar controls derived from engineered plants that overproduce lignin-related aromatics. Here this enhancement was more pronounced (~seven-fold increase) with hydrolysates from nonengineered switchgrass. Proteomic analysis of the methyl ketone-producing P. putida suggested that plant-derived amino acids may be the source of this enhancement. Mass spectrometry-based measurements of plant-derived amino acids demonstrated a high correlation between methyl ketone production and amino acid concentration in plant hydrolysates. Amendment of glucose-containing minimal media with a defined mixture of amino acids similar to those found in the hydrolysates studied led to a nine-fold increase in methyl ketone titer (1.1 g/L).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Amino Acid Immobilization of Copper Surface Diffusion on Cu(111)

Surface diffusion and molecular self-assembly are two critically important processes in chemistry and nature. Amino acids deposited on a Cu(111) surface driving a separation at the 2D limit between self-assembling molecules and diffusing copper atoms is reported. Since the self-assembling amino acids prefer non-planar, tridentate bonding with neighboring adatoms, they attach to and immobilize diffusing copper adatoms on the surface. This chemical interaction freezes out the copper diffusion causing the condensation of solid copper adatom islands on the surface. Such separation and immobilization are observed for eight different amino acids, suggesting the generality of this phenomenon beyond a single amino acid species. Furthermore, at elevated temperatures, a disruption of the prototypical Ostwald ripening of adatom islands is also observed. Here, these results provide fundamental insight into chiral molecular self-assembly and its interplay with metal atom surface diffusion.

36 MATERIALS SCIENCE↗
DOE OSTI2018

Biochemical Characterization of β-Amino Acid Incorporation in Fluvirucin B 2 Biosynthesis

Naturally occurring lactams, such as the polyketide-derived macrolactams, provide a diverse class of natural products that could enhance existing chemically produced lactams. While β-amino acid loading in the fluvirucin B 2 polyketide pathway has been proposed by a previously identified putative biosynthetic gene cluster, biochemical characterization of the complete loading enzymes has not been described. In this paper, we elucidate the complete biosynthetic pathway of the β-amino acid loading pathway in fluvirucin B 2 biosynthesis. We demonstrate the promiscuity of the loading pathway to utilize a range of amino acids and further illustrate the ability to introduce non-native acyl transferases to selectively transfer β-amino acids onto a PKS loading platform. The results presented here provide a detailed biochemical description of β-amino acid selection and will further aid in future efforts to develop engineered lactam-producing PKS platforms.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

Dual incorporation of non-canonical amino acids enables production of post-translationally modified selenoproteins

Post-translational modifications (PTMs) can occur on almost all amino acids in eukaryotes as a key mechanism for regulating protein function. The ability to study the role of these modifications in various biological processes requires techniques to modify proteins site-specifically. One strategy for this is genetic code expansion (GCE) in bacteria. The low frequency of post-translational modifications in bacteria makes it a preferred host to study whether the presence of a post-translational modification influences a protein’s function. Genetic code expansion employs orthogonal translation systems engineered to incorporate a modified amino acid at a designated protein position. Selenoproteins, proteins containing selenocysteine, are also known to be post-translationally modified. Selenoproteins have essential roles in oxidative stress, immune response, cell maintenance, and skeletal muscle regeneration. Their complicated biosynthesis mechanism has been a hurdle in our understanding of selenoprotein functions. As technologies for selenocysteine insertion have recently improved, we wanted to create a genetic system that would allow the study of post-translational modifications in selenoproteins. By combining genetic code expansion techniques and selenocysteine insertion technologies, we were able to recode stop codons for insertion of N ε -acetyl- l -lysine and selenocysteine, respectively, into multiple proteins. The specificity of these amino acids for their assigned position and the simplicity of reverting the modified amino acid via mutagenesis of the codon sequence demonstrates the capacity of this method to study selenoproteins and the role of their post-translational modifications. Moreover, the evidence that Sec insertion technology can be combined with genetic code expansion tools further expands the chemical biology applications.

59 BASIC BIOLOGICAL SCIENCES↗