Construction of carotenoid biosynthetic pathways using squalene synthase

Furubayashi, Maiko; Li, Ling; Katabami, Akinori; Saito, Kyoichi; Umeno, Daisuke

doi:10.1016/j.febslet.2013.12.003

Cited by 33 publications

(38 citation statements)

References 33 publications

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“…In S. aureus , biosynthesis of the carotenoid staphyloxanthin is catalyzed by the dehydrosqualene synthase CrtM, which converts farnesyl diphosphate (FPP) to dehydrosqualene (31). A recent study has determined that combining the activities of SQS and dehydrosqualene desaturase (CrtN) was sufficient to synthesize the carotenoid pigment staphyloxanthin by combining E. coli and S. aureus enzymes, respectively (31). Experimentation is required to determine catalytic specificity of the putative Staphylococcus group B-associated SQS.…”

Section: Resultsmentioning

confidence: 99%

“…SQS catalyzes an alternative pathway for the biosynthesis of carotenoid pigments (30). In S. aureus , biosynthesis of the carotenoid staphyloxanthin is catalyzed by the dehydrosqualene synthase CrtM, which converts farnesyl diphosphate (FPP) to dehydrosqualene (31). A recent study has determined that combining the activities of SQS and dehydrosqualene desaturase (CrtN) was sufficient to synthesize the carotenoid pigment staphyloxanthin by combining E. coli and S. aureus enzymes, respectively (31).…”

Section: Resultsmentioning

confidence: 99%

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Comparative Genomics of Staphylococcus Reveals Determinants of Speciation and Diversification of Antimicrobial Defense

Horsburgh

Coates-Brown

Darby

et al. 2018

Preprint

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26The bacterial genus Staphylococcus comprises diverse species with most being described 27 as colonizers of human and animal skin. A relational analysis of features that 28 discriminate its species and contribute to niche adaptation and survival remains to be fully 29 described. In this study, an interspecies, whole-genome comparative analysis of 21 30Staphylococcus species was performed based on their orthologues. Three well-defined 31 multi-species groups were identified: group A (including aureus/epidermidis); group B 32 (including saprophyticus/xylosus) and group C (including pseudintermedius/delphini). 33The machine learning algorithm Random Forest was applied to identify variable 34 orthologues that drive formation of the Staphylococcus species groups A-C. Orthologues 35 driving staphylococcal infrageneric diversity comprised regulatory, metabolic and 36 antimicrobial resistance proteins. Notably, the BraSR (NsaRS) two-component system 37 (TCS) and its associated BraDE transporters that regulate antimicrobial resistance 38 distinguish group A Staphylococcus species from others in the genus that lack the BraSR 39 TCS. Divergence of BraSR and GraSR antimicrobial peptide survival TCS and their 40 associated transporters was observed across the staphylococci, likely reflecting niche 41 specific evolution of these TCS/transporters and their specificities for AMPs. 42 Experimental evolution, with selection for resistance to the lantibiotic nisin, revealed 43 multiple routes to resistance and differences in the selection outcomes of the BraSR-44 positive species S. hominis and S. aureus. Selection supported a role for GraSR in nisin 45 survival responses of the BraSR-negative group B species S. saprophyticus. Our study 46 reveals diversification of antimicrobial-sensing TCS across the staphylococci and hints at 47 differential relationships between GraSR and BraSR in those species positive for both 48 TCS. 49 50 Importance 51The genus Staphylococcus includes species that are commensals and opportunist 52 pathogens of humans and animals. Identifying the features that discriminate species of 53 staphylococci is relevant to understanding niche selection and the structure of their 54 microbiomes. Moreover, the determinants that structure the community are relevant for 55 strategies to modify the frequency of individual species associated with dysbiosis and 56 disease. In this study, we identify orthologous proteins that discriminate genomes of 57 staphylococci. In particular, species restriction of a major antimicrobial survival system, 58 BraSR (NsaRS), to a group of staphylococci dominated by those that can colonize human 59 skin. The diversity of antimicrobial sensing loci was revealed by comparative analysis 60 3 and experimental evolution with selection for nisin resistance identified the potential for 61 variation in antimicrobial sensing in BraRS-encoding staphylococci. This study provides 62 insights into staphylococcal species diversity. 63 64 Introduction 65 Staphylococcus species and genomics 66The existen...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Comparative Genomics of Staphylococcus Reveals Determinants of Speciation and Diversification of Antimicrobial Defense

Horsburgh

Coates-Brown

Darby

et al. 2018

Preprint

View full text Add to dashboard Cite

show abstract

“…2) was analyzed as previously described [51] with a slight modification. Plasmids (TPS genes on the pUC vector and carotenoid genes on the pAC vector) were transformed into XL1-Blue cells, and the transformants were plated onto LB-Lennox (carb/cm) agar plates to form colonies.…”

Section: Methodsmentioning

confidence: 99%

A High-Throughput Colorimetric Screening Assay for Terpene Synthase Activity Based on Substrate Consumption

et al. 2014

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Terpene synthases catalyze the formation of a variety of terpene chemical structures. Systematic mutagenesis studies have been effective in providing insights into the characteristic and complex mechanisms of C-C bond formations and in exploring the enzymatic potential for inventing new chemical structures. In addition, there is growing demand to increase terpene synthase activity in heterologous hosts, given the maturation of metabolic engineering and host breeding for terpenoid synthesis. We have developed a simple screening method for the cellular activities of terpene synthases by scoring their substrate consumption based on the color loss of the cell harboring carotenoid pathways. We demonstrate that this method can be used to detect activities of various terpene synthase or prenyltransferase genes in a high-throughput manner, irrespective of the product type, enabling the mutation analysis and directed evolution of terpene synthases. We also report the possibility for substrate-specific screening system of terpene synthases by taking advantage of the substrate-size specificity of C30 and C40 carotenoid pathways.

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“…pAC‐ hsqs and pUCara‐ crtI were derived from Ref. [20]. Here, the hsqs gene was derived from pHSS16 from Thompson et al [28], where 30 N‐terminal residues and 47 C‐terminal residues were truncated.…”

Section: Methodsmentioning

confidence: 99%

“…CrtM catalyzes the same first reaction, but the second rearrangement step produces DSQ instead of SQ. CrtN catalyzes the desaturation of DSQ, also using SQ as a substrate [20] to produce diaponeurosporene and/or diapolycopene, C 30 carotenoid pigments. CrtI, conversely, uses DSQ but not SQ.…”

Section: Introductionmentioning

confidence: 99%

Directed evolution of squalene synthase for dehydrosqualene biosynthesis

Furubayashi

Katabami

et al. 2014

FEBS Letters

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a b s t r a c tSqualene synthase (SQS) catalyzes the first step of sterol/hopanoid biosynthesis in various organisms. It has been long recognized that SQSs share a common ancestor with carotenoid synthases, but it is not known how these enzymes selectively produce their own product. In this study, SQSs from yeast, human, and bacteria were independently subjected to directed evolution for the production of the C 30 carotenoid backbone, dehydrosqualene. This was accomplished via high-throughput screening with Pantoea ananatis phytoene desaturase, which can selectively convert dehydrosqualene into yellow carotenoid pigments. Genetic analysis of the resultant mutants revealed various mutations that could effectively convert SQS into a ''dehydrosqualene synthase.'' All of these mutations are clustered around the residues that have been proposed to be important for NADPH binding.

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Construction of carotenoid biosynthetic pathways using squalene synthase

Cited by 33 publications

References 33 publications

Comparative Genomics of Staphylococcus Reveals Determinants of Speciation and Diversification of Antimicrobial Defense

Comparative Genomics of Staphylococcus Reveals Determinants of Speciation and Diversification of Antimicrobial Defense

A High-Throughput Colorimetric Screening Assay for Terpene Synthase Activity Based on Substrate Consumption

Directed evolution of squalene synthase for dehydrosqualene biosynthesis

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