Fig 5 - uploaded by Tobias Erb
Content may be subject to copyright.
Proposed pathway for the assimilation of C1 compounds by isocitrate lyase-negative type II methylotrophs by using the serine cycle for formaldehyde fixation. The ethylmalonyl-CoA pathway described here (excluding the condensation of acetyl-CoA and glyoxylate) is integrated in the serine cycle (upper part) and is involved in assimilation of acetyl-CoA and regeneration of glyoxylate during growth on C 1 compounds. It is assumed that during growth on C 2 compounds the ethylmalonyl-CoA pathway (Fig. 4) is used exclusively. Dotted lines indicate more than one reaction step.
Source publication
Fifty years ago, Kornberg and Krebs established the glyoxylate cycle as the pathway for the synthesis of cell constituents from C2-units. However, since then, many bacteria have been described that do not contain isocitrate lyase, the key enzyme of this pathway. Here, a pathway termed the ethylmalonyl-CoA pathway operating in such organisms is desc...
Contexts in source publication
Context 1
... of acetyl-CoA to glyoxylate (6,19). Here, we have shown high crotonyl-CoA carboxylase/reductase activity in cell extracts of methanol-grown M. extorquens. The presence of the ethylmalonyl-CoA pathway (excluding the condensation of acetyl-CoA and glyoxylate) in M. extorquens simplifies the scheme for isocitrate lyase-negative C 1 - assimilation (Fig. 5) compared with former proposals (12). Genes of the ethylmalonyl-CoA pathway, including ccr, were affected in mutants of M. extorquens unable to form glyoxylate from acetyl-CoA. However, different catalytic roles were ini- tially assigned to the corresponding gene products (12,16,20). It has been reported that marine species of the ...
Context 2
... 1 -compounds by these aerobic phototrophs has not been explicitly shown. All genes implicated in the herein-described ethylmalonyl-CoA pathway, including ccr, now known to encode the key enzyme crotonyl-CoA carboxylase/ reductase, are conserved in sequenced representatives of this phylogenetic group (13), suggesting that the extended serine cycle (Fig. 5) may be involved in metabolism of further ecologically important compounds such as ...
Similar publications
The glyoxylate cycle is an anaplerotic pathway of the tricarboxylic acid (TCA) cycle that allows growth on C(2) compounds by bypassing the CO(2)-generating steps of the TCA cycle. The unique enzymes of this route are isocitrate lyase (ICL) and malate synthase (MS). ICL cleaves isocitrate to glyoxylate and succinate, and MS converts glyoxylate and a...
Citations
... Therefore, the type of electron carrier used is a key difference between the assimilation of acetate and fructose. The term NADH dehydrogenase (ubiquinone) activity (GO:0008137) could refer to the initial step in the electron transport chain [54,55] and reflect this difference in the assimilation pathway. This change in electron delivery to the membrane seems to also be found in the term electron transfer activity (GO:0009055). ...
Purple photosynthetic bacteria (PPB) are versatile microorganisms capable of producing various value-added chemicals, e.g., biopolymers and biofuels. They employ diverse metabolic pathways, allowing them to adapt to various growth conditions and even extreme environments. Thus, they are ideal organisms for the Next Generation Industrial Biotechnology concept of reducing the risk of contamination by using naturally robust extremophiles. Unfortunately, the potential of PPB for use in biotechnology is hampered by missing knowledge on regulations of their metabolism. Although Rhodospirillum rubrum represents a model purple bacterium studied for polyhydroxyalkanoate and hydrogen production, light/chemical energy conversion, and nitrogen fixation, little is known regarding the regulation of its metabolism at the transcriptomic level. Using RNA sequencing, we compared gene expression during the cultivation utilizing fructose and acetate as substrates in case of the wild-type strain R. rubrum DSM 467T and its knock-out mutant strain that is missing two polyhydroxyalkanoate synthases PhaC1 and PhaC2. During this first genome-wide expression study of R. rubrum, we were able to characterize cultivation-driven transcriptomic changes and to annotate non-coding elements as small RNAs.
... Recently, a new class of CO 2 -fixing enzymes, so-called enoyl-CoA carboxylases/reductases (ECRs), were discovered in alphaproteobacteria, where they serve in central acetate metabolism. These ECRs catalyze the NADPH-dependent reductive carboxylation of enoyl-CoA thioester into their corresponding alkyl-malonyl-CoA esters (Erb et al. 2007). ECRs are particularly interesting, because they capture CO 2 at up to 10fold higher rates compared to rubisco (i.e., up to 100 CO 2 molecules per second). ...
... ECRs are particularly interesting, because they capture CO 2 at up to 10fold higher rates compared to rubisco (i.e., up to 100 CO 2 molecules per second). Moreover, they do not show any oxygenase side reaction (Erb et al. 2007(Erb et al. , 2009DeMirci et al. 2022). Overall, this makes ECRs an interesting alternative to rubisco-based CO 2 -fixation. ...
... The CETCH cycle is a network of 17 enzymes that is thermodynamically and kinetically favored compared to the Calvin cycle of natural photosynthesis. The CETCH cycle requires 20% less energy and the carboxylation rates of crotonyl-CoA and acrylyl-CoA by ECR outcompete rubisco by almost one order of magnitude with respect to CO 2 -fixation rates (Erb et al. 2007;Schwander et al. 2016;DeMirci et al. 2022). ...
Synthetic biology provides opportunities to realize new-to-nature CO2-fixation metabolisms to overcome the limitations of natural photosynthesis. Two different strategies are currently being pursued: One is to realize engineered plants that feature carbon-neutral or carbon-negative (i.e., CO2-fixing) photorespiration metabolism, such as the tatronyl-CoA (TaCo) pathway, to boost CO2-uptake rates of photosynthesis between 20% and 60%. Another (arguably more radical) is to create engineered plants in which natural photosynthesis is fully replaced by an alternative CO2-fixation metabolism, such as the CETCH cycle, which carries the potential to improve CO2 uptake rates between 20% and 200%. These efforts could revolutionize plant engineering by expanding the capabilities of plant metabolism beyond the constraints of natural evolution to create highly improved crops addressing the challenges of climate change in the future.
... 017510335 (alignment fraction = 0.747). S2 is predicted to contain the ethylmalonyl pathway, which has been used as a way to utilize acetate in bacteria lacking isocitrate lyase (19). S2 contains SoxBCDYZ, Sqr, and FccB and is capable of thiosulfate oxidation to sulfate in vitro (6). ...
Here, we report the complete genome sequence for Roseinatronobacter sp. S2, a sulfur-oxidizing heterotroph isolated from a serpentinizing system in Northern California. The S2 genome is 4.4 Mb and contains 4,570 protein-encoding genes. This organism contains the genes necessary for sulfur species oxidation and complete ethylmalonyl and pentose phosphate pathways.
... Several classes of enzymes important to bioelectrosynthesis exhibit NAD-dependent behavior including dehydrogenases and reductases, [73,75] reductases. [76][77] Unfortunately, both NAD and NADPH exhibit poor electrochemical reversibility, being prone to dimerization to inactive species upon direct reduction at an electrode. [57,78] Thus a cofactor regeneration system must be employed to catalytic turnover in these systems. ...
Bioelectrocatalytic synthesis is the conversion of electrical energy into value‐added products using biocatalysts. These methods merge the specificity and selectivity of biocatalysis and energy‐related electrocatalysis to address challenges in the sustainable synthesis of pharmaceuticals, commodity chemicals, fuels, feedstocks and fertilizers. However, the specialized experimental setups and domain knowledge for bioelectrocatalysis pose a significant barrier to adoption. This review introduces key concepts of bioelectrosynthetic systems. We provide a tutorial on the methods of biocatalyst utilization, the setup of bioelectrosynthetic cells, and the analytical methods for assessing bioelectrocatalysts. Key applications of bioelectrosynthesis in ammonia production and small‐molecule synthesis are outlined for both enzymatic and microbial systems. This review serves as a necessary introduction and resource for the non‐specialist interested in bioelectrosynthetic research.
... Several classes of enzymes important to bioelectrosynthesis exhibit NAD-dependent behavior including dehydrogenases and reductases, [73,75] reductases. [76][77] Unfortunately, both NAD and NADPH exhibit poor electrochemical reversibility, being prone to dimerization to inactive species upon direct reduction at an electrode. [57,78] Thus a cofactor regeneration system must be employed to catalytic turnover in these systems. ...
Bioelectrocatalytic synthesis is the conversion of electrical energy into value-added chemicals via a biocatalyst. Bioelectrosynthetic methods utilize the specificity and selectivity of biocatalytic enzymatic or microbial species to carry out chemical redox transformations while utilizing electricity as a stoichiometric redox equivalent. As a merging of biocatalysis and electrocatalysis, these methods directly address challenges in green and sustainable synthesis of pharmaceuticals, commodity chemicals, fuels, feedstocks and fertilizers. Despite the rising importance of bioelectrochemical transformations across these industries, there remains a high barrier for adoption due to the specialized experimental setups and domain knowledge for bioelectrocatalysis. This review aims to introduce the key concepts and design features of bioelectrosynthetic systems. A tutorial on the methods of biocatalyst utilization and the setup of bioelectrosynthetic cells is provided, as well as an overview of the analytical methods used for assessing bioelectrocatalysts. Key studies illustrating the vital applications of bioelectrosynthesis are outlined, such as ammonia production, small-molecule synthesis, and multi-carbon product formation. Finally, we address future directions for both microbial and enzymatic electrosynthetic methods. In summary, this review provides a critically necessary introduction to the field and a collection of resources for the non-specialist interested in pursuing a research program in bioelectrosynthesis.
... While acetyl-CoA is oxidized through the tricarboxylic acid (TCA) cycle for NADH and ATP generation, bacteria have evolved different strategies for the assimilation of acetyl-CoA into biomass. Several acetyl-CoA assimilation routes have been described to date (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12) among them the glyoxylate cycle (2,3) and the ethylmalonyl-CoA pathway (4)(5)(6)(7)12). The glyoxylate cycle (GC) consists of only two enzymes, isocitrate lyase (Icl) and malate synthase (Ms), which branch off the TCA cycle and are often organized in one operon under joint transcriptional regulation (13). ...
... While acetyl-CoA is oxidized through the tricarboxylic acid (TCA) cycle for NADH and ATP generation, bacteria have evolved different strategies for the assimilation of acetyl-CoA into biomass. Several acetyl-CoA assimilation routes have been described to date (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12) among them the glyoxylate cycle (2,3) and the ethylmalonyl-CoA pathway (4)(5)(6)(7)12). The glyoxylate cycle (GC) consists of only two enzymes, isocitrate lyase (Icl) and malate synthase (Ms), which branch off the TCA cycle and are often organized in one operon under joint transcriptional regulation (13). ...
... The ethylmalonyl-CoA pathway (EMCP), instead, is a rather complex pathway that involves at least 12 enzymes (4), among them crotonyl-CoA carboxylase/ reductase (Ccr). Ccr catalyzes a unique reaction, the reductive carboxylation of crotonyl-CoA into ethylmalonyl-CoA, and is considered the key enzyme of the EMCP (4,6). In contrast to the GC, genes encoding the EMCP are typically distributed across the host genome (14). ...
Carbon metabolism provides organisms with energy and building blocks for cellular functions and growth. The tight regulation between degradation and assimilation of carbon substrates is central for optimal growth.
... Different routes to generate biosynthetic precursors from acetate assimilation also determine the balance of electrons in PNSB. R. sphaeroids uses the reductive ethylmalonyl-CoA pathway, while R. palustris uses the oxidative glyoxylate shunt (154,159). In R. sphaeroids which relies on Rnf for Fd production, the reductive ethylmalonyl-CoA pathway consumes reductant in the initial assimilation of acetate to malate and negates the use of the CBB cycle as an electron sink (159). ...
... R. sphaeroids uses the reductive ethylmalonyl-CoA pathway, while R. palustris uses the oxidative glyoxylate shunt (154,159). In R. sphaeroids which relies on Rnf for Fd production, the reductive ethylmalonyl-CoA pathway consumes reductant in the initial assimilation of acetate to malate and negates the use of the CBB cycle as an electron sink (159). In contrast, R. palustris, which relies on the Fix complex for Fd production, uses the glyoxylate shunt to produce net NADH and requires an electron sink (160). ...
The availability of fixed nitrogen is a limiting factor in the net primary production of all ecosystems. Diazotrophs overcome this limit through the conversion of atmospheric dinitrogen to ammonia. Diazotrophs are phylogenetically diverse bacteria and archaea that exhibit a wide range of lifestyles and metabolisms, including obligate anaerobes and aerobes that generate energy through heterotrophic or autotrophic metabolisms. Despite the diversity of metabolisms, all diazotrophs use the same enzyme, nitrogenase, to reduce N2. Nitrogenase is an O2-sensitive enzyme that requires a high amount of energy in the form of ATP and low potential electrons carried by ferredoxin (Fd) or flavodoxin (Fld). This review summarizes how the diverse metabolisms of diazotrophs utilize different enzymes to generate low potential reducing equivalents for nitrogenase catalysis. These enzymes include substrate-level Fd oxidoreductases, hydrogenases, photosystem I or other light-driven reaction centers, electron bifurcating Fix complexes, proton motive force-driven Rnf complexes, and Fd:NAD(P)H oxidoreductases. Each of these enzymes is critical for generating low potential electrons while simultaneously integrating the native metabolism to balance nitrogenase's overall energy needs. Understanding the diversity of electron transport systems to nitrogenase in various diazotrophs will be essential to guide future engineering strategies aimed at expanding the contributions of biological nitrogen fixation in agriculture.
... CETCH循环是第一条非天然人工 固碳途径, 包含来自古菌、细菌、植物等9种生物的17 种酶, 并在几轮酶工程中得到优化 [45,47] . 其核心固碳酶 巴豆酰CoA羧化酶/还原酶(Ccr)是迄今为止固碳活性最 高的羧化酶, 与RuBisco和其他羧化酶相比, Ccr具有广 泛的底物谱, 对氧不敏感, 催化CO 2 的固定效率比Ru-bisCO高2~4倍, 但在进化过程中并没有用于自养固碳 途径 [48] . 通过CETCH将CO 2 转化为有机产物的体外代 谢过程优于化学过程, CETCH的固碳效率达到5 nmol 表 1 6种人工固碳途径的比较 [49] . ...
... 一般 来说, 因为在厌氧条件下没有氧气, 因此电极产生较少 腐蚀, 故更有利于电能自养过程 [89] . [50] , 例如PEP羧化酶 [58] 和crotonyl-CoA 羧化酶 [105] 等新酶的发现以及通过定向进化改造RuBis-CO [106] 从而提高碳捕获酶的催化性能. 此外, 值得关注 的其他挑战还有: 目前可应用的自养生物通常细胞生 长缓慢、产物合成能力低、难以基因修饰等问题 [4] . ...
... Unlike the two former pathways mentioned above, here formaldehyde is first converted to formate and then to 5, 10-methylene-tetrahydrofolate (CH2-THF) before incorporating into the pathway using tetrahydrofolate (THF) as the cofactor (Crowther et al., 2008). CH2-THF condenses with glycine (derived from glyoxylate) to form serine; however, (unlike the regeneration of C5-P in RuMP and XuMP via PPP pathway) regeneration of glyoxylate occurs through ethylmalonyl-CoA (EMC) pathway circulating the Serine cycle (Erb et al., 2007;Peyraud et al., 2009). While operating the serine cycle, an isotopic labeling study confirmed the incorporation of one CO2, along with the incorporation of each formaldehyde, to form acetyl-CoA (Crowther et al., 2008), pointing toward a dual mode of carbon assimilation. ...
Methanol, a relatively cheap and renewable single-carbon feedstock, has gained considerable attention as a substrate for the bio-production of commodity chemicals. Conventionally produced from syngas, along with emerging possibilities of generation from methane and CO2, this C1 substrate can serve as a pool for sequestering greenhouse gases while supporting a sustainable bio-economy. Methylotrophic organisms, with the inherent ability to use methanol as the sole carbon and energy source, are competent candidates as platform organisms. Accordingly, methanol bioconversion pathways have been an attractive target for biotechnological and bioengineering interventions in developing microbial cell factories. This review summarizes the recent advances in methanol-based production of various bulk and value-added chemicals exploiting the native and synthetic methylotrophic organisms. Finally, the current challenges and prospects of streamlining these methylotrophic platforms are discussed.
