Molecular cloning and characterization of the ATP citrate lyase from carotenogenic yeast<i>Phaffia rhodozyma</i>

Chávez-Cabrera, Cipriano; Marsch, Rodolfo; Bartolo-Aguilar, Yaneth; Flores-Bustamante, Zoila R.; Hidalgo-Lara, María Eugenia; Martínez-Cárdenas, Anahí; Cancino‐Díaz, Juan C.; Sánchez, Sergio; Flores-Cotera, Luis B.

doi:10.1093/femsyr/fov054

Cited by 4 publications

(4 citation statements)

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“…ACL is present in all eukaryotes except non-oleaginous yeasts. In animals and oleaginous basidiomycete yeasts, ACL is encoded by a single gene [26,27]; in plants and some filamentous fungi, ACL usually consists of two subunits (ACL1 and ACL2) with homology to the N-and C-terminals of the animal ATPcitrate lyase polypeptide [28]. Fatty acid content was increased in Y. lipolytica by overexpression of ACL1 and ACL2 on a non-lipogenic medium in an obese strain [29] or overexpression of ACL from Mus musculus [30].…”

Section: Introductionmentioning

confidence: 99%

Enhancement of docosahexaenoic acid production by overexpression of ATP-citrate lyase and acetyl-CoA carboxylase in Schizochytrium sp.

Xiao

Zhao

Wen

et al. 2020

Biotechnol Biofuels

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Background: Docosahexaenoic acid (DHA) is an important omega-3 long-chain polyunsaturated fatty acid that has a variety of physiological functions for infant development and human health. Although metabolic engineering was previously demonstrated to be a highly efficient way to rapidly increase lipid production, metabolic engineering has seldom been previously used to increase DHA accumulation in Schizochytrium spp. Results: Here, a sensitive β-galactosidase reporter system was established to screen for strong promoters in Schizochytrium sp. Four constitutive promoters (EF-1α p , TEF-1 p , ccg1 p , and ubiquitin p) and one methanol-induced AOX1 promoter were characterized by the reporter system with the promoter activity ccg1 p > TEF-1 p > AOX1 p (induced) > EF-1α p > ubiquitin p. With the strong constitutive promoter ccg1 p , Schizochytrium ATP-citrate lyase (ACL) and acetyl-CoA carboxylase (ACC) were overexpressed in Schizochytrium sp. ATCC 20888. The cells were cultivated at 28 °C and 250 rpm for 120 h with glucose as the carbon source. Shake-flask fermentation results showed that the overexpression strains exhibited growth curves and biomass similar to those of the wild-type strain. The lipid contents of the wild-type strain and of the OACL, OACC, and OACL-ACC strains were 53.8, 68.8, 69.8, and 73.0%, respectively, and the lipid yields of the overexpression strains were increased by 21.9, 30.5, and 38.3%, respectively. DHA yields of the wild-type strain and of the corresponding overexpression strains were 4.3, 5.3, 6.1, and 6.4 g/L, i.e., DHA yields of the overexpression strains were increased by 23.3, 41.9, and 48.8%, respectively. Conclusions: Acetyl-CoA and malonyl-CoA are precursors for fatty acid synthesis. ACL catalyzes the conversion of citrate in the cytoplasm into acetyl-CoA, and ACC catalyzes the synthesis of malonyl-CoA from acetyl-CoA. The results demonstrate that overexpression of ACL and ACC enhances lipid accumulation and DHA production in Schizochytrium sp.

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Section: Introductionmentioning

confidence: 99%

Enhancement of docosahexaenoic acid production by overexpression of ATP-citrate lyase and acetyl-CoA carboxylase in Schizochytrium sp.

Xiao

Zhao

Wen

et al. 2020

Biotechnol Biofuels

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“…Nitrogen depletion may cause the reduction of ACL activity, thereby repressing AX production [25]. As hypothesized by Chávez-Cabrera et al, there may be a second isoform of ACL that affects the balance between lipid and carotenoid synthesis and drives the metabolism pathway toward lipid over carotenoid production upon nitrogen deficiency, thereby reducing AX production [25,26]. In addition, the C/N ratio can cause differences in the proteomics profile.…”

Section: Astaxanthin Production With Meat and Bone Meal Hydrolysates ...mentioning

confidence: 96%

Utilization of Meat and Bone Meal for Yeast Fermentation to Produce Astaxanthin

Zheng

2023

Fermentation

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Meat and bone meal (MBM) is a product of the rendering industry, which is looking for high-value applications of rendered animal proteins (RAP). The objective of this research was to utilize MBM as a nitrogen source to produce astaxanthin (AX) by Xanthophyllomyces dendrorhous and quantify the bioavailability of MBM as a potential substitution of commercial nitrogen sources (i.e., yeast extract and peptone). To conduct yeast fermentation under the optimal glucose loading, the C/N ratio was optimized to achieve maximum AX content. MBM was hydrolyzed by using proteinase and alkaline (Ca(OH)2) for 4, 8, and 16 h with different enzyme and alkaline loadings to produce MBM hydrolysates (MBMHs). The MBMHs were directly fermented by X. dendrorhous under the optimum glucose concentration. Experimentally, the optimum medium contained 40 g/L glucose, 5 g/L peptone, and 3 g/L yeast extract, where AX content of 3.69 mg/g dry cell mass was achieved. MBMHs were used by X. dendrorhous as a nitrogen source, while fermentation with lyophilized MBMHs was generated using proteinase K. This resulted in a maximum AX content of 1.58 mg/g dry cell mass. This research exhibits the feasibility of using MBM as a nitrogen source to produce AX with X. dendrorhous.

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“…Some studies have shown that low-temperature expression of recombinant proteins from diverse origins (prokaryotes to higher eukaryotes), in P. pastoris, can increase the yield, activity, stability, secretion, and solubility [168][169][170][171][172][173][174]. For example, heterologous expression of Vitreoscilla hemoglobin (VHb) improves cell growth and recombinant protein production in various hosts, including P. pastoris.…”

Section: Cold-responsive For Protein Expression In Yeastsmentioning

confidence: 99%

“…However, the active form of the enzyme expressed was only detected when the recombinant yeast was grown at 25 °C and 300 rpm. The authors suggested that the production of the active recombinant enzyme may demand post-translational modifications such as phosphorylation and/or a suitable folding [168,177]. Some other studies have reported successful expression at low temperatures in S. cerevisiae, S. pombe, Kluyveromyces marxianus, and Y. lipolytica [178][179][180][181][182].…”

Section: Cold-responsive For Protein Expression In Yeastsmentioning

confidence: 99%

The potential of cold-shock promoters for the expression of recombinant proteins in microbes and mammalian cells

Bartolo-Aguilar

Chávez-Cabrera

Flores-Cotera

et al. 2022

Journal of Genetic Engineering and Biotechnology

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Background Low-temperature expression of recombinant proteins may be advantageous to support their proper folding and preserve bioactivity. The generation of expression vectors regulated under cold conditions can improve the expression of some target proteins that are difficult to express in different expression systems. Main body of the abstract The cspA encodes the major cold-shock protein from Escherichia coli (CspA). The promoter of cspA has been widely used to develop cold shock-inducible expression platforms in E. coli. Moreover, it is often necessary to employ expression systems other than bacteria, particularly when recombinant proteins require complex post-translational modifications. Currently, there are no commercial platforms available for expressing target genes by cold shock in eukaryotic cells. Consequently, genetic elements that respond to cold shock offer the possibility of developing novel cold-inducible expression platforms, particularly suitable for yeasts, and mammalian cells. Conclusions This review covers the importance of the cellular response to low temperatures and the prospective use of cold-sensitive promoters to direct the expression of recombinant proteins. This concept may contribute to renewing interest in applying white technologies to produce recombinant proteins that are difficult to express. Graphical Abstract

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Molecular cloning and characterization of the ATP citrate lyase from carotenogenic yeastPhaffia rhodozyma

Cited by 4 publications

References 36 publications

Enhancement of docosahexaenoic acid production by overexpression of ATP-citrate lyase and acetyl-CoA carboxylase in Schizochytrium sp.

Enhancement of docosahexaenoic acid production by overexpression of ATP-citrate lyase and acetyl-CoA carboxylase in Schizochytrium sp.

Utilization of Meat and Bone Meal for Yeast Fermentation to Produce Astaxanthin

The potential of cold-shock promoters for the expression of recombinant proteins in microbes and mammalian cells

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