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![Fatty acid and phosphatidic acid synthesis in Bacillales. Malonyl‐CoA is generated from acetyl‐CoA by acetyl‐CoA carboxylase (ACC), and then, is transferred to ACP by malonyl‐CoA transacylase (FabD). The malonate group enters the type II fatty acid synthase (FASII), to generate an acyl‐ACP two carbons longer than the original acyl‐ACP at the end of each cycle. When the acyl chains reach an appropriate length, they become substrates of the acyl‐transferase enzymes to generate phosphatidic acid (PA). First, PlsX catalyzes the synthesis of acyl‐phosphate (acyl‐PO4) from acyl‐ACP; then, PlsY transfers the fatty acid from acyl‐PO4 to the 1‐position of glycerol‐3‐phosphate (G3P). Finally, lyso‐PA (LPA) is acylated by PlsC obtaining PA. Free fatty acids (FFAs) can be phosphorylated by the fatty acid kinase FakA/FakB. The resultant acyl‐PO4s can be either incorporated into the 1‐position of G3P by PlsY or converted to acyl‐ACP by PlsX to be elongated by FASII or utilized by PlsC. Malonyl‐CoA and malonyl‐ACP inhibit the transcriptional repressor FapR, which controls at least 10 genes (the fap regulon) coding for FASII enzymes, PlsX and PlsC [Colour figure can be viewed at wileyonlinelibrary.com]](publication/342985988/figure/fig1/AS:11431281176517513@1690194259166/Fatty-acid-and-phosphatidic-acid-synthesis-in-Bacillales-Malonyl-CoA-is-generated-from.png)
Fatty acid and phosphatidic acid synthesis in Bacillales. Malonyl‐CoA is generated from acetyl‐CoA by acetyl‐CoA carboxylase (ACC), and then, is transferred to ACP by malonyl‐CoA transacylase (FabD). The malonate group enters the type II fatty acid synthase (FASII), to generate an acyl‐ACP two carbons longer than the original acyl‐ACP at the end of each cycle. When the acyl chains reach an appropriate length, they become substrates of the acyl‐transferase enzymes to generate phosphatidic acid (PA). First, PlsX catalyzes the synthesis of acyl‐phosphate (acyl‐PO4) from acyl‐ACP; then, PlsY transfers the fatty acid from acyl‐PO4 to the 1‐position of glycerol‐3‐phosphate (G3P). Finally, lyso‐PA (LPA) is acylated by PlsC obtaining PA. Free fatty acids (FFAs) can be phosphorylated by the fatty acid kinase FakA/FakB. The resultant acyl‐PO4s can be either incorporated into the 1‐position of G3P by PlsY or converted to acyl‐ACP by PlsX to be elongated by FASII or utilized by PlsC. Malonyl‐CoA and malonyl‐ACP inhibit the transcriptional repressor FapR, which controls at least 10 genes (the fap regulon) coding for FASII enzymes, PlsX and PlsC [Colour figure can be viewed at wileyonlinelibrary.com]
Source publication
A key aspect in membrane biogenesis is the coordination of fatty acid to phospholipid synthesis rates. In most bacteria, PlsX is the first enzyme of the phosphatidic acid synthesis pathway, the common precursor of all phospholipids. Previously, we proposed that PlsX is a key regulatory point that synchronizes the fatty acid synthase II with phospho...
Citations
... For example, in B. subtilis, FAS and phospholipid synthesis are coordinated in response to the availability of the malonyl-CoA precursor, which binds as an inducer to the FapR repressor (30). Depletion of the fatty acyl-phosphate synthase, PlsX, halts both FAS and phospholipid synthesis, which suggests that plsX also plays a key regulatory role in coordinating FAS and phospholipid synthesis (42,43). Similarly, PG synthesis is homeostatically regulated by multiple mechanisms that control the GlmS-dependent partition of fructose 6-phosphate into amino sugar synthesis. ...
Proper synthesis and maintenance of a multilayered cell envelope are critical for bacterial fitness. However, whether mechanisms exist to coordinate synthesis of the membrane and peptidoglycan layers is unclear. In Bacillus subtilis, synthesis of peptidoglycan (PG) during cell elongation is mediated by an elongasome complex acting in concert with class A penicillin-binding proteins (aPBPs). We previously described mutant strains limited in their capacity for PG synthesis due to a loss of aPBPs and an inability to compensate by upregulation of elongasome function. Growth of these PG-limited cells can be restored by suppressor mutations predicted to decrease membrane synthesis. One suppressor mutation leads to an altered function repressor, FapR*, that functions as a super-repressor and leads to decreased transcription of fatty acid synthesis (FAS) genes. Consistent with fatty acid limitation mitigating cell wall synthesis defects, inhibition of FAS by cerulenin also restored growth of PG-limited cells. Moreover, cerulenin can counteract the inhibitory effect of β-lactams in some strains. These results imply that limiting PG synthesis results in impaired growth, in part, due to an imbalance of PG and cell membrane synthesis and that B. subtilis lacks a robust physiological mechanism to reduce membrane synthesis when PG synthesis is impaired. IMPORTANCE Understanding how a bacterium coordinates cell envelope synthesis is essential to fully appreciate how bacteria grow, divide, and resist cell envelope stresses, such as β-lactam antibiotics. Balanced synthesis of the peptidoglycan cell wall and the cell membrane is critical for cells to maintain shape and turgor pressure and to resist external cell envelope threats. Using Bacillus subtilis, we show that cells deficient in peptidoglycan synthesis can be rescued by compensatory mutations that decrease the synthesis of fatty acids. Further, we show that inhibiting fatty acid synthesis with cerulenin is sufficient to restore growth of cells deficient in peptidoglycan synthesis. Understanding the coordination of cell wall and membrane synthesis may provide insights relevant to antimicrobial treatment.
... FAs are synthesized from acetyl-ACP and malonyl-ACP, catalyzed by the FA synthase complex, through condensation, reduction, dehydration, and reduction reactions. The first and rate-limiting step in the initiation of fatty acid biosynthesis is mediated by acetyl-CoA carboxylase (ACC), which catalyzes the production of malonyl-CoA from acetyl-ACP (Machinandiarena et al. 2020). As shown in Table 4, accA was downregulated 2.07-fold, indicating that the FA biosynthesis process was limited at pH 4.0. ...
Streptomyces albulus is a well-established cell factory for ε-poly-L-lysine (ε-PL) production. It has been reported that ε-PL biosynthesis is strictly regulated by pH and that ε-PL can accumulate at approximately pH 4.0, which is outside of the general pH range for natural product production by Streptomyces species. However, how S. albulus responds to low pH is not clear. In this study, we attempted to explore the response of S. albulus to low-pH stress at the physiological and global gene transcription levels. At the physiological level, S. albulus maintained intracellular pH homeostasis at ~pH 7.5, increased the unsaturated fatty acid ratio, extended the fatty acid chain length, enhanced ATP accumulation, increased H⁺-ATPase activity, and accumulated the basic amino acids L-lysine and L-arginine. At the global gene transcription level, carbohydrate metabolism, oxidative phosphorylation, macromolecule protection and repair, and the acid tolerance system were found to be involved in combating low-pH stress. Finally, we preliminarily evaluated the effect of the acid tolerance system and cell membrane fatty acid synthesis on low-pH tolerance via gene manipulation. This work provides new insight into the adaptation mechanism of Streptomyces to low-pH stress and a new opportunity for constructing robust S. albulus strains for ε-PL production.
Key points
• S. albulus consistently remained pHiat ~7.4 regardless of the environmental pH.
• S. albulus combats low-pH stress by modulating lipid composition of cell membrane.
• Overexpression of cfa in S. albulus could improve low-pH tolerance and ɛ-PL titer.
Graphical abstract
... Fatty acids are used for fueling the citrate cycle and the glyoxylate shunt for generating energy via the fatty acid degradation pathway [19]. Fatty acid chains also form membrane phospholipids through the glycerolipid and glycerphospholipid biosynthesis pathways, thus playing a critical role in membrane biogenesis [20,21]. Thus, MFFabG4 Functional interaction of MFFabG4 to identify interacting proteins was accomplished using a homologous protein FabG4 of M. tuberculosis H37Rv as M. ...
The fatty acid biosynthesis pathway is crucial for the formation of the mycobacterial cell envelope. The fatty acid synthase type‐II (FAS‐II) components are attractive targets for designing anti‐biofilm inhibitors. Literature review, bioinformatics analysis, cloning, and sequencing led to the identification of a novel Mycobacterium fortuitum FAS‐II gene MFfabG4 which interacts with mycobacterial proteins involved in biofilm formation. A manually curated M. fortuitum fatty acid biosynthesis pathway has been proposed exploiting functional studies from the Kyoto Encyclopedia of Genes and Genomes and Mycobrowser databases for MFFabG4. M. fortuitum MFfabG4 knockdown strain (FA) was constructed and validated by quantitative polymerase chain reaction. The FA strain displayed unstructured smooth colony architecture, correlating with decreased pathogenicity and virulence. MFfabG4 knockdown resulted in diminished pellicle and attenuated biofilm formation, along with impaired sliding motility, and reduced cell sedimentation. The FA strain showed lowered cell surface hydrophobicity, indicating attenuation in M. fortuitum intracellular infection‐causing ability. Stress survival studies showed the requirement of MFfabG4 for survival in a nutrient‐starved environment. The results indicate that MFfabG4 maintains the physiology of the cell envelope and is required for the formation of M. fortuitum pellicle and biofilm. The study corroborates the role of MFfabG4 as a pellicle‐ and biofilm‐specific drug target and a potential diagnostic marker for M. fortuitum and related pathogenic mycobacteria.
... FapR is the key transcriptional regulator of membrane lipid synthesis in B. subtilis and clinically relevant pathogens such as Staphylococcus aureus, Bacillus anthracis, and Listeria monocytogenes (Schujman et al., 2003;Fujita et al., 2007;Albanesi et al., 2013;Machinandiarena et al., 2020), and modulates the overall rate of membrane synthesis in response to precursor availability. B. subtilis FapR represses genes for FA and phospholipid synthesis, and this repression is relieved by allosteric interactions with malonyl-CoA or malonyl-ACP (Schujman et al., 2006;Martinez et al., 2010). ...
... As a branchpoint enzyme, ACC is often under complex regulation (Zhang and Rock, 2009;Salie and Thelen, 2016;Machinandiarena et al., 2020). In B. subtilis, ACC is regulated in part by YqhY, a conserved DUF322/Asp23 protein which is highly expressed and often encoded together with ACC subunits as part of an accB-accC-yqhY operon (Todter et al., 2017). ...
Antibiotics and other agents that perturb the synthesis or integrity of the bacterial cell envelope trigger compensatory stress responses. Focusing on Bacillus subtilis as a model system, this mini-review summarizes current views of membrane structure and insights into how cell envelope stress responses remodel and protect the membrane. Altering the composition and properties of the membrane and its associated proteome can protect cells against detergents, antimicrobial peptides, and pore-forming compounds while also, indirectly, contributing to resistance against compounds that affect cell wall synthesis. Many of these regulatory responses are broadly conserved, even where the details of regulation may differ, and can be important in the emergence of antibiotic resistance in clinical settings.
Fatty acid biosynthesis (FASII) enzymes are considered valid targets for antimicrobial drug development against the human pathogen Staphylococcus aureus . However, incorporation of host fatty acids confers FASII antibiotic adaptation that compromises prospective treatments. S. aureus adapts to FASII inhibitors by first entering a nonreplicative latency period, followed by outgrowth. Here, we used transcriptional fusions and direct metabolite measurements to investigate the factors that dictate the duration of latency prior to outgrowth. We show that stringent response induction leads to repression of FASII and phospholipid synthesis genes. (p)ppGpp induction inhibits synthesis of malonyl-CoA, a molecule that derepresses FapR, a key regulator of FASII and phospholipid synthesis. Anti-FASII treatment also triggers transient expression of (p)ppGpp-regulated genes during the anti-FASII latency phase, with concomitant repression of FapR regulon expression. These effects are reversed upon outgrowth. GTP depletion, a known consequence of the stringent response, also occurs during FASII latency, and is proposed as the common signal linking these responses. We next showed that anti-FASII treatment shifts malonyl-CoA distribution between its interactants FapR and FabD, toward FapR, increasing expression of the phospholipid synthesis genes plsX and plsC during outgrowth. We conclude that components of the stringent response dictate malonyl-CoA availability in S. aureus FASII regulation, and contribute to latency prior to anti-FASII-adapted outgrowth. A combinatory approach, coupling a (p)ppGpp inducer and an anti-FASII, blocks S. aureus outgrowth, opening perspectives for bi-therapy treatment.
IMPORTANCE Staphylococcus aureus is a major human bacterial pathogen for which new inhibitors are urgently needed. Antibiotic development has centered on the fatty acid synthesis (FASII) pathway, which provides the building blocks for bacterial membrane phospholipids. However, S. aureus overcomes FASII inhibition and adapts to anti-FASII by using exogenous fatty acids that are abundant in host environments. This adaptation mechanism comprises a transient latency period followed by bacterial outgrowth. Here, we use metabolite sensors and promoter reporters to show that responses to stringent conditions and to FASII inhibition intersect, in that both involve GTP and malonyl-CoA. These two signaling molecules contribute to modulating the duration of latency prior to S. aureus adaptation outgrowth. We exploit these novel findings to propose a bi-therapy treatment against staphylococcal infections.
