@phdthesis{Moratti2020,
  author    = {Moratti, Fabio Giulio},
  title     = {Structural analysis of DYW proteins and identification of the mitochondrial DNA-binding proteome of Arabidopsis thaliana},
  school      = {Universit{\"a}t Potsdam},
  pages     = {164},
  year      = {2020},
  language  = {en}
}
@phdthesis{Naake2020,
  author    = {Naake, Thomas},
  title     = {Strategies to investigate the natural variation of plant specialized metabolism},
  school      = {Universit{\"a}t Potsdam},
  year      = {2020},
  language  = {en}
}
@phdthesis{Nowak2020,
  author    = {Nowak, Jacqueline},
  title     = {Devising computational tools to quantify the actin cytoskeleton and pavement cell shape using network-based approaches},
  school      = {Universit{\"a}t Potsdam},
  pages     = {123},
  year      = {2020},
  abstract  = {Recent advances in microscopy have led to an improved visualization of different cell processes. Yet, this also leads to a higher demand of tools which can process images in an automated and quantitative fashion. Here, we present two applications that were developed to quantify different processes in eukaryotic cells which rely on the organization and dynamics of the cytoskeleton.. In plant cells, microtubules and actin filaments form the backbone of the cytoskeleton. These structures support cytoplasmic streaming, cell wall organization and tracking of cellular material to and from the plasma membrane. To better understand the underlying mechanisms of cytoskeletal organization, dynamics and coordination, frameworks for the quantification are needed. While this is fairly well established for the microtubules, the actin cytoskeleton has remained difficult to study due to its highly dynamic behaviour. One aim of this thesis was therefore to provide an automated framework to quantify and describe actin organization and dynamics. We used the framework to represent actin structures as networks and examined the transport efficiency in Arabidopsis thaliana hypocotyl cells. Furthermore, we applied the framework to determine the growth mode of cotton fibers and compared the actin organization in wild-type and mutant cells of rice. Finally, we developed a graphical user interface for easy usage. Microtubules and the actin cytoskeleton also play a major role in the morphogenesis of epidermal leaf pavement cells. These cells have highly complex and interdigitated shapes which are hard to describe in a quantitative way. While the relationship between microtubules, the actin cytoskeleton and shape formation is the object of many studies, it is still not clear how and if the cytoskeletal components predefine indentations and protrusions in pavement cell shapes. To understand the underlying cell processes which coordinate cell morphogenesis, a quantitative shape descriptor is needed. Therefore, the second aim of this thesis was the development of a network-based shape descriptor which captures global and local shape features, facilitates shape comparison and can be used to evaluate shape complexity. We demonstrated that our framework can be used to describe and compare shapes from various domains. In addition, we showed that the framework accurately detects local shape features of pavement cells and outperform contending approaches. In the third part of the thesis, we extended the shape description framework to describe pavement cell shape features on tissue-level by proposing different network representations of the underlying imaging data.},
  language  = {en}
}
@phdthesis{Saplaoura2020,
  author    = {Saplaoura, Eleftheria},
  title     = {Escaping the plant cell},
  school      = {Universit{\"a}t Potsdam},
  pages     = {156},
  year      = {2020},
  language  = {en}
}
@phdthesis{Schuster2020,
  author    = {Schuster, Maja},
  title     = {High resolution decoding of the tobacco chloroplast translatome and its dynamics during light-intensity acclimation},
  doi       = {10.25932/publishup-51268},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-512680},
  school      = {Universit{\"a}t Potsdam},
  pages     = {xvii, 155},
  year      = {2020},
  abstract  = {Chloroplasts are the photosynthetic organelles in plant and algae cells that enable photoautotrophic growth. Due to their prokaryotic origin, modern-day chloroplast genomes harbor 100 to 200 genes. These genes encode for core components of the photosynthetic complexes and the chloroplast gene expression machinery, making most of them essential for the viability of the organism. The regulation of those genes is predominated by translational adjustments. The powerful technique of ribosome profiling was successfully used to generate highly resolved pictures of the translational landscape of Arabidopsis thaliana cytosol, identifying translation of upstream open reading frames and long non-coding transcripts. In addition, differences in plastidial translation and ribosomal pausing sites were addressed with this method. However, a highly resolved picture of the chloroplast translatome is missing. Here, with the use of chloroplast isolation and targeted ribosome affinity purification, I generated highly enriched ribosome profiling datasets of the chloroplasts translatome for Nicotiana tabacum in the dark and light. Chloroplast isolation was found unsuitable for the unbiased analysis of translation in the chloroplast but adequate to identify potential co-translational import. Affinity purification was performed for the small and large ribosomal subunit independently. The enriched datasets mirrored the results obtained from whole-cell ribosome profiling. Enhanced translational activity was detected for psbA in the light. An alternative translation initiation mechanism was not identified by selective enrichment of small ribosomal subunit footprints. In sum, this is the first study that used enrichment strategies to obtain high-depth ribosome profiling datasets of chloroplasts to study ribosome subunit distribution and chloroplast associated translation. Ever-changing light intensities are challenging the photosynthetic capacity of photosynthetic organism. Increased light intensities may lead to over-excitation of photosynthetic reaction centers resulting in damage of the photosystem core subunits. Additional to an expensive repair mechanism for the photosystem II core protein D1, photosynthetic organisms developed various features to reduce or prevent photodamage. In the long-term, photosynthetic complex contents are adjusted for the efficient use of experienced irradiation. However, the contribution of chloroplastic gene expression in the acclimation process remained largely unknown. Here, comparative transcriptome and ribosome profiling was performed for the early time points of high-light acclimation in Nicotiana tabacum chloroplasts in a genome-wide scale. The time- course data revealed stable transcript level and only minor changes in translational activity of specific chloroplast genes during high-light acclimation. Yet, psbA translation was increased by two-fold in the high light from shortly after the shift until the end of the experiment. A stress-inducing shift from low- to high light exhibited increased translation only of psbA. This study indicate that acclimation fails to start in the observed time frame and only short-term responses to reduce photoinhibition were observed.},
  language  = {en}
}
@phdthesis{Schaelicke2020,
  author    = {Sch{\"a}licke, Svenja},
  title     = {Consumer traits and trait variation under the influence of biochemical food quality},
  school      = {Universit{\"a}t Potsdam},
  pages     = {136},
  year      = {2020},
  abstract  = {The earth's ecosystems undergo considerable changes characterized by human-induced alterations of environmental factors. In order to develop conservation goals for vulnerable ecosystems, research on ecosystem functioning is required.. Therefore, it is crucial to explore organismal interactions, such as trophic interaction or competition, which are decisive for key processes in ecosystems. These interactions are determined by the performance responses of organisms to environmental changes, which in turn, are shaped by the organism's functional traits. Exploring traits, their variation, and the environmental factors that act on them may provide insights on how ecological interactions affect populations, community structures and dynamics, and thus ecosystem functioning. In aquatic ecosystems, global warming intensifies phytoplankton blooms, which are more frequently dominated by cyanobacteria. As cyanobacteria are poor in polyunsaturated fatty acids (PUFA) and sterols, this compositional change alters the biochemical food quality of phytoplankton for consumer species with potential effects on ecological interactions. Within this thesis, I studied the effects of biochemical food quality on consumer traits and performance responses at the phytoplankton-zooplankton interface using different strains of two closely related generalist rotifer species Brachionus calyciflorus and Brachionus fernandoi and three phytoplankton species that differ in their biochemical food quality, i.e. in their content and composition of PUFA and sterols. In a series of laboratory feeding experiments I found that biochemical food quality affected rotifer's performance, i.e. fecundity, survival, and population growth, across a broad range of food quantities. Biochemical food quality constraints, which are often underestimated as influencing environmental factors, had strong impacts on performance responses. I further explored the potential of biochemical food quality in mediating consumer response variation between species and among strains of one species. Co-limitation by food quantity and biochemical food quality resulted in differences in performance responses, which were more pronounced within than between rotifer species. Furthermore, I demonstrated that the body PUFA compositions of rotifer species and strains were differently affected by the dietary PUFA supply, which indicates inter- and intraspecific differences in physiological traits, such as PUFA retention, allocation, and/or bioconversion capacity, within the genus Brachionus. This indicates that dietary PUFA are involved in shaping traits and performance responses of rotifers. This thesis reveals that biochemical food quality is an environmental factor with strong effects on individual traits and performance responses of consumers. Biochemical food quality constraints can further mediate trait and response variation among species or strains. Consequently, they carry the potential to shape ecological interactions and evolutionary processes with effects on community structures and dynamics. Trait-based approaches, which include food quality research, thus may provide further insights into the linkage between functional diversity and the maintenance of crucial ecosystem functions.},
  language  = {en}
}
@phdthesis{Siemiatkowska2020,
  author    = {Siemiatkowska, Beata},
  title     = {Redox signalling in plants},
  doi       = {10.25932/publishup-48911},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-489119},
  school      = {Universit{\"a}t Potsdam},
  pages     = {127},
  year      = {2020},
  abstract  = {Once proteins are synthesized, they can additionally be modified by post-translational modifications (PTMs). Proteins containing reactive cysteine thiols, stabilized in their deprotonated form due to their local environment as thiolates (RS-), serve as redox sensors by undergoing a multitude of oxidative PTMs (Ox-PTMs). Ox-PTMs such as S-nitrosylation or formation of inter- or intra-disulfide bridges induce functional changes in these proteins. Proteins containing cysteines, whose thiol oxidation state regulates their functions, belong to the so-called redoxome. Such Ox-PTMs are controlled by site-specific cellular events that play a crucial role in protein regulation, affecting enzyme catalytic sites, ligand binding affinity, protein-protein interactions or protein stability. Reversible protein thiol oxidation is an essential regulatory mechanism of photosynthesis, metabolism, and gene expression in all photosynthetic organisms. Therefore, studying PTMs will remain crucial for understanding plant adaptation to external stimuli like fluctuating light conditions. Optimizing methods suitable for studying plants Ox-PTMs is of high importance for elucidation of the redoxome in plants. This study focusses on thiol modifications occurring in plant and provides novel insight into in vivo redoxome of Arabidopsis thaliana in response to light vs. dark. This was achieved by utilizing a resin-assisted thiol enrichment approach. Furthermore, confirmation of candidates on the single protein level was carried out by a differential labelling approach. The thiols and disulfides were differentially labelled, and the protein levels were detected using immunoblot analysis. Further analysis was focused on light-reduced proteins. By the enrichment approach many well studied redox-regulated proteins were identified. Amongst those were fructose 1,6-bisphosphatase (FBPase) and sedoheptulose-1,7-bisphosphatase (SBPase) which have previously been described as thioredoxin system targeted enzymes. The redox regulated proteins identified in the current study were compared to several published, independent results showing redox regulated proteins in Arabidopsis leaves, root, mitochondria and specifically S-nitrosylated proteins. These proteins were excluded as potential new candidates but remain as a proof-of-concept to the enrichment experiments to be effective. Additionally, CSP41A and CSP41B proteins, which emerged from this study as potential targets of redox-regulation, were analyzed by Ribo-Seq. The active translatome study of csp41a mutant vs. wild-type showed most of the significant changes at end of the night, similarly as csp41b. Yet, in both mutants only several chloroplast-encoded genes were altered. Further studies of CSP41A and CSP41B proteins are needed to reveal their functions and elucidate the role of redox regulation of these proteins.},
  language  = {en}
}
@phdthesis{Tunn2020,
  author    = {Tunn, Isabell},
  title     = {From single molecules to bulk materials: tuning the viscoelastic properties of coiled coil cross-linked hydrogels},
  doi       = {10.25932/publishup-47595},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-475955},
  school      = {Universit{\"a}t Potsdam},
  pages     = {XVI, 140},
  year      = {2020},
  abstract  = {The development of bioinspired self-assembling materials, such as hydrogels, with promising applications in cell culture, tissue engineering and drug delivery is a current focus in material science. Biogenic or bioinspired proteins and peptides are frequently used as versatile building blocks for extracellular matrix (ECM) mimicking hydrogels. However, precisely controlling and reversibly tuning the properties of these building blocks and the resulting hydrogels remains challenging. Precise control over the viscoelastic properties and self-healing abilities of hydrogels are key factors for developing intelligent materials to investigate cell matrix interactions. Thus, there is a need to develop building blocks that are self-healing, tunable and self-reporting. This thesis aims at the development of α-helical peptide building blocks, called coiled coils (CCs), which integrate these desired properties. Self-healing is a direct result of the fast self-assembly of these building blocks when used as material cross-links. Tunability is realized by means of reversible histidine (His)-metal coordination bonds. Lastly, implementing a fluorescent readout, which indicates the CC assembly state, self-reporting hydrogels are obtained. Coiled coils are abundant protein folding motifs in Nature, which often have mechanical function, such as in myosin or fibrin. Coiled coils are superhelices made up of two or more α-helices wound around each other. The assembly of CCs is based on their repetitive sequence of seven amino acids, so-called heptads (abcdefg). Hydrophobic amino acids in the a and d position of each heptad form the core of the CC, while charged amino acids in the e and g position form ionic interactions. The solvent-exposed positions b, c and f are excellent targets for modifications since they are more variable. His-metal coordination bonds are strong, yet reversible interactions formed between the amino acid histidine and transition metal ions (e.g. Ni2+, Cu2+ or Zn2+). His-metal coordination bonds essentially contribute to the mechanical stability of various high-performance proteinaceous materials, such as spider fangs, Nereis worm jaws and mussel byssal threads. Therefore, I bioengineered reversible His-metal coordination sites into a well-characterized heterodimeric CC that served as tunable material cross-link. Specifically, I took two distinct approaches facilitating either intramolecular (Chapter 4.2) and/or intermolecular (Chapter 4.3) His-metal coordination. Previous research suggested that force-induced CC unfolding in shear geometry starts from the points of force application. In order to tune the stability of a heterodimeric CC in shear geometry, I inserted His in the b and f position at the termini of force application (Chapter 4.2). The spacing of His is such that intra-CC His-metal coordination bonds can form to bridge one helical turn within the same helix, but also inter-CC coordination bonds are not generally excluded. Starting with Ni2+ ions, Raman spectroscopy showed that the CC maintained its helical structure and the His residues were able to coordinate Ni2+. Circular dichroism (CD) spectroscopy revealed that the melting temperature of the CC increased by 4 °C in the presence of Ni2+. Using atomic force microscope (AFM)-based single molecule force spectroscopy, the energy landscape parameters of the CC were characterized in the absence and the presence of Ni2+. His-Ni2+ coordination increased the rupture force by ~10 pN, accompanied by a decrease of the dissociation rate constant. To test if this stabilizing effect can be transferred from the single molecule level to the bulk viscoelastic material properties, the CC building block was used as a non-covalent cross-link for star-shaped poly(ethylene glycol) (star-PEG) hydrogels. Shear rheology revealed a 3-fold higher relaxation time in His-Ni2+ coordinating hydrogels compared to the hydrogel without metal ions. This stabilizing effect was fully reversible when using an excess of the metal chelator ethylenediaminetetraacetate (EDTA). The hydrogel properties were further investigated using different metal ions, i.e. Cu2+, Co2+ and Zn2+. Overall, these results suggest that Ni2+, Cu2+ and Co2+ primarily form intra-CC coordination bonds while Zn2+ also participates in inter-CC coordination bonds. This may be a direct result of its different coordination geometry. Intermolecular His-metal coordination bonds in the terminal regions of the protein building blocks of mussel byssal threads are primarily formed by Zn2+ and were found to be intimately linked to higher-order assembly and self-healing of the thread. In the above example, the contribution of intra-CC and inter-CC His-Zn2+ cannot be disentangled. In Chapter 4.3, I redesigned the CC to prohibit the formation of intra-CC His-Zn2+ coordination bonds, focusing only on inter-CC interactions. Specifically, I inserted His in the solvent-exposed f positions of the CC to focus on the effect of metal-induced higher-order assembly of CC cross-links. Raman and CD spectroscopy revealed that this CC building block forms α-helical Zn2+ cross-linked aggregates. Using this CC as a cross-link for star-PEG hydrogels, I showed that the material properties can be switched from viscoelastic in the absence of Zn2+ to elastic-like in the presence of Zn2+. Moreover, the relaxation time of the hydrogel was tunable over three orders of magnitude when using different Zn2+:His ratios. This tunability is attributed to a progressive transformation of single CC cross-links into His-Zn2+ cross-linked aggregates, with inter-CC His-Zn2+ coordination bonds serving as an additional, cross-linking mode. Rheological characterization of the hydrogels with inter-CC His-Zn2+ coordination raised the question whether the His-Zn2+ coordination bonds between CCs or also the CCs themselves rupture when shear strain is applied. In general, the amount of CC cross-links initially formed in the hydrogel as well as the amount of CC cross-links breaking under force remains to be elucidated. In order to more deeply probe these questions and monitor the state of the CC cross-links when force is applied, a fluorescent reporter system based on F{\"o}rster resonance energy transfer (FRET) was introduced into the CC (Chapter 4.4). For this purpose, the donor-acceptor pair carboxyfluorescein and tetramethylrhodamine was used. The resulting self-reporting CC showed a FRET efficiency of 77 \% in solution. Using this fluorescently labeled CC as a self-reporting, reversible cross-link in an otherwise covalently cross-linked star-PEG hydrogel enabled the detection of the FRET efficiency change under compression force. This proof-of-principle result sets the stage for implementing the fluorescently labeled CCs as molecular force sensors in non-covalently cross-linked hydrogels. In summary, this thesis highlights that rationally designed CCs are excellent reversibly tunable, self-healing and self-reporting hydrogel cross-links with high application potential in bioengineering and biomedicine. For the first time, I demonstrated that His-metal coordination-based stabilization can be transferred from the single CC level to the bulk material with clear viscoelastic consequences. Insertion of His in specific sequence positions was used to implement a second non-covalent cross-linking mode via intermolecular His-metal coordination. This His-metal binding induced aggregation of the CCs enabled for reversibly tuning the hydrogel properties from viscoelastic to elastic-like. As a proof-of-principle to establish self-reporting CCs as material cross-links, I labeled a CC with a FRET pair. The fluorescently labelled CC acts as a molecular force sensor and first preliminary results suggest that the CC enables the detection of hydrogel cross-link failure under compression force. In the future, fluorescently labeled CC force sensors will likely not only be used as intelligent cross-links to study the failure of hydrogels but also to investigate cell-matrix interactions in 3D down to the single molecule level.},
  language  = {en}
}
@phdthesis{Wen2020,
  author    = {Wen, Xi},
  title     = {Distribution patterns and environmental drivers of methane-cycling microorganisms in natural environments and restored wetlands},
  doi       = {10.25932/publishup-47177},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-471770},
  school      = {Universit{\"a}t Potsdam},
  pages     = {VIII, iii, 152},
  year      = {2020},
  abstract  = {Methane is an important greenhouse gas contributing to global climate change. Natural environments and restored wetlands contribute a large proportion to the global methane budget. Methanogenic archaea (methanogens) and methane oxidizing bacteria (methanotrophs), the biogenic producers and consumers of methane, play key roles in the methane cycle in those environments. A large number of studies revealed the distribution, diversity and composition of these microorganisms in individual habitats. However, uncertainties exist in predicting the response and feedback of methane-cycling microorganisms to future climate changes and related environmental changes due to the limited spatial scales considered so far, and due to a poor recognition of the biogeography of these important microorganisms combining global and local scales. With the aim of improving our understanding about whether and how methane-cycling microbial communities will be affected by a series of dynamic environmental factors in response to climate change, this PhD thesis investigates the biogeographic patterns of methane-cycling communities, and the driving factors which define these patterns at different spatial scales. At the global scale, a meta-analysis was performed by implementing 94 globally distributed public datasets together with environmental data from various natural environments including soils, lake sediments, estuaries, marine sediments, hydrothermal sediments and mud volcanos. In combination with a global biogeographic map of methanogenic archaea from multiple natural environments, this thesis revealed that biogeographic patterns of methanogens exist. The terrestrial habitats showed higher alpha diversities than marine environments. Methanoculleus and Methanosaeta (Methanothrix) are the most frequently detected taxa in marine habitats, while Methanoregula prevails in terrestrial habitats. Estuary ecosystems, the transition zones between marine and terrestrial/limnic ecosystems, have the highest methanogenic richness but comparably low methane emission rates. At the local scale, this study compared two rewetted fens with known high methane emissions in northeastern Germany, a coastal brackish fen (H{\"u}telmoor) and a freshwater riparian fen (Polder Zarnekow). Consistent with different geochemical conditions and land-use history, the two rewetted fens exhibit dissimilar methanogenic and, especially, methanotrophic community compositions. The methanotrophic community was generally under-represented among the prokaryotic communities and both fens show similarly low ratios of methanotrophic to methanogenic abundances. Since few studies have characterized methane-cycling microorganisms in rewetted fens, this study provides first evidence that the rapid and well re-established methanogenic community in combination with the low and incomplete re-establishment of the methanotrophic community after rewetting contributes to elevated sustained methane fluxes following rewetting. Finally, this thesis demonstrates that dispersal limitation only slightly regulates the biogeographic distribution patterns of methanogenic microorganisms in natural environments and restored wetlands. Instead, their existence, adaption and establishment are more associated with the selective pressures under different environmental conditions. Salinity, pH and temperature are identified as the most important factors in shaping microbial community structure at different spatial scales (global versus terrestrial environments). Predicted changes in climate, such as increasing temperature, changes in precipitation patterns and increasing frequency of flooding events, are likely to induce a series of environmental alterations, which will either directly or indirectly affect the driving environmental forces of methanogenic communities, leading to changes in their community composition and thus potentially also in methane emission patterns in the future.},
  language  = {en}
}
@phdthesis{Wenk2020,
  author    = {Wenk, Sebastian},
  title     = {Engineering formatotrophic growth in Escherichia coli},
  school      = {Universit{\"a}t Potsdam},
  pages     = {V, 107},
  year      = {2020},
  abstract  = {To meet the demands of a growing world population while reducing carbon dioxide (CO2) emissions, it is necessary to capture CO2 and convert it into value-added compounds. In recent years, metabolic engineering of microbes has gained strong momentum as a strategy for the production of valuable chemicals. As common microbial feedstocks like glucose directly compete with human consumption, the one carbon (C1) compound formate was suggested as an alternative feedstock. Formate can be easily produced by various means including electrochemical reduction of CO2 and could serve as a feedstock for microbial production, hence presenting a novel entry point for CO2 to the biosphere and a storage option for excess electricity. Compared to the gaseous molecule CO2, formate is a highly soluble compound that can be easily handled and stored. It can serve as a carbon and energy source for natural formatotrophs, but these microbes are difficult to cultivate and engineer. In this work, I present the results of several projects that aim to establish efficient formatotrophic growth of E. coli - which cannot naturally grow on formate - via synthetic formate assimilation pathways. In the first study, I establish a workflow for growth-coupled metabolic engineering of E. coli. I demonstrate this approach by presenting an engineering scheme for the PFL-threonine cycle, a synthetic pathway for anaerobic formate assimilation in E. coli. The described methods are intended to create a standardized toolbox for engineers that aim to establish novel metabolic routes in E. coli and related organisms. The second chapter presents a study on the catalytic efficiency of C1-oxidizing enzymes in vivo. As formatotrophic growth requires generation of both energy and biomass from formate, the engineered E. coli strains need to be equipped with a highly efficient formate dehydrogenase, which provides reduction equivalents and ATP for formate assimilation. I engineered a strain that cannot generate reducing power and energy for cellular growth, when fed on acetate. Under this condition, the strain depends on the introduction of an enzymatic system for NADH regeneration, which could further produce ATP via oxidative phosphorylation. I show that the strain presents a valuable testing platform for C1-oxidizing enzymes by testing different NAD-dependent formate and methanol dehydrogenases in the energy auxotroph strain. Using this platform, several candidate enzymes with high in vivo activity, were identified and characterized as potential energy-generating systems for synthetic formatotrophic or methylotrophic growth in E. coli.   In the third chapter, I present the establishment of the serine threonine cycle (STC) - a synthetic formate assimilation pathway - in E. coli. In this pathway, formate is assimilated via formate tetrahydrofolate ligase (FtfL) from Methylobacterium extorquens (M. extorquens). The carbon from formate is attached to glycine to produce serine, which is converted into pyruvate entering central metabolism. Via the natural threonine synthesis and cleavage route, glycine is regenerated and acetyl-CoA is produced as the pathway product. I engineered several selection strains that depend on different STC modules for growth and determined key enzymes that enable high flux through threonine synthesis and cleavage. I could show that expression of an auxiliary formate dehydrogenase was required to achieve growth via threonine synthesis and cleavage on pyruvate. By overexpressing most of the pathway enzymes from the genome, and applying adaptive laboratory evolution, growth on glycine and formate was achieved, indicating the activity of the complete cycle. The fourth chapter shows the establishment of the reductive glycine pathway (rGP) - a short, linear formate assimilation route - in E. coli. As in the STC, formate is assimilated via M. extorquens FtfL. The C1 from formate is condensed with CO2 via the reverse reaction of the glycine cleavage system to produce glycine. Another carbon from formate is attached to glycine to form serine, which is assimilated into central metabolism via pyruvate. The engineered E. coli strain, expressing most of the pathway genes from the genome, can grow via the rGP with formate or methanol as a sole carbon and energy source.},
  language  = {en}
}