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Jae Hyung Lee Yong Bae Seo Seong-Yun Jeong Soo-Wan Nam Young Tae Kim 《Biotechnology and Bioprocess Engineering》2007,12(3):312-317
Carotenoids are important natural pigments produced by many microorganisms and plants. We have previously reported the isolation
of a new marine bacterium,Paracoccus haeundaensis, which produces carotenoids, mainly in the form of astaxanthin. The astaxanthin biosynthesis gene cluster, consisting of
six carotenogenic genes, was cloned and characterized from this organism. Individual genes of the carotenoid biosynthesis
gene cluster were functionally expressed inEscherichia coli and each gene product was purified to homogeneity. Their molecular characteristics, including enzymatic activities, were
previously reported. Here, we report cloning the genes for crtE, crtEB, crtEBI, crtEBIY, crtEBIYZ, and crtEBI-YZW of theP. haeundaensis carotenoid biosynthesis genes inE. coli and verifying the production of the corresponding pathway intermediates. The carotenoids that accumulated in the transformed
cells carrying these gene combinations were analyzed by chromatographic and spectroscopic methods. 相似文献
3.
Carotenoids are indispensable pigments of the photosynthetic apparatus in plants, algae, and cyanobacteria and are produced,
as well, by many bacteria and fungi. Elucidation of biochemical pathways leading to the carotenoids that function in the photosynthetic
membranes of land plants has been greatly aided by the use of carotenoid-accumulating strains of Escherichia coli as heterologous hosts for functional assays, in vivo, of the otherwise difficult to study membrane-associated pathway enzymes.
This same experimental approach is uniquely well-suited to the discovery and characterization of yet-to-be identified enzymes
that lead to carotenoids of the photosynthetic membranes in algal cells, to the multitude of carotenoids found in nongreen
plant tissues, and to the myriad flavor and aroma compounds that are derived from carotenoids in plant tissues. A portfolio
of plasmids suitable for the production in E. coli of a variety of carotenoids is presented herein. The use of these carotenoid-producing E. coli for the identification of cDNAs encoding enzymes of carotenoid and isoprenoid biosynthesis, for characterization of the enzymes
these cDNAs encode, and for the production of specific carotenoids for use as enzyme substrates and reference standards, is
described using the flowering plant Adonis aestivalis to provide examples. cDNAs encoding nine different A. aestivalis enzymes of carotenoid and isoprenoid synthesis were identified and the enzymatic activity of their products verified. Those
cDNAs newly described include ones that encode phytoene synthase, β-carotene hydroxylase, deoxyxylulose-5-phosphate synthase,
isopentenyl diphosphate isomerase, and geranylgeranyl diphosphate synthase. 相似文献
4.
Carotenoids represent a group of valuable molecules for the pharmaceutical, chemical, food and feed industries, not only because
they can act as vitamin A precursors, but also for their coloring, antioxidant and possible tumor-inhibiting activity. Animals
cannot synthesize carotenoids, and these pigments must therefore be added to the feeds of farmed species. The synthesis of
different natural commercially important carotenoids (β-carotene, torulene, torularhodin and astaxanthin) by several yeast
species belonging to the genera Rhodotorula and Phaffia has led to consider these microorganisms as a potential pigment sources. In this review, we discuss the biosynthesis, factors
affecting carotenogenesis in Rhodotorula and Phaffia strains, strategies for improving the production properties of the strains and directions for potential utility of carotenoid-synthesizing
yeast as a alternative source of natural carotenoid pigments. 相似文献
5.
J. Wu B.A. Boghigian M. Myint H. Zhang S. Zhang B.A. Pfeifer 《Letters in applied microbiology》2010,51(2):196-204
Aims: Escherichia coli has emerged as a viable heterologous host for the production of complex, polyketide natural compounds. In this study, polyketide biosynthesis was compared between different E. coli strains for the purpose of better understanding and improving heterologous production. Methods and Results: Both B and K‐12 E. coli strains were genetically modified to support heterologous polyketide biosynthesis [specifically, 6‐deoxyerythronolide B (6dEB)]. Polyketide production was analysed using a helper plasmid designed to overcome rare codon usage within E. coli. Each strain was analysed for recombinant protein production, precursor consumption, by‐product production, and 6dEB biosynthesis. Of the strains tested for biosynthesis, 6dEB production was greatest for E. coli B strains. When comparing biosynthetic improvements as a function of mRNA stability vs codon bias, increased 6dEB titres were observed when additional rare codon tRNA molecules were provided. Conclusions: Escherichia coli B strains and the use of tRNA supplementation led to improved 6dEB polyketide titres. Significance and Impact of the Study: Given the medicinal potential and growing field of polyketide heterologous biosynthesis, the current study provides insight into host‐specific genetic backgrounds and gene expression parameters aiding polyketide production through E. coli. 相似文献
6.
The recently discovered non-mevalonate pathway to isoprenoids, which uses glycolytic intermediates, has been modulated by
overexpression of Escherichia coli
d-1-deoxyxylulose 5-phosphate synthase (DXS) to increase deoxyxylulose 5-phosphate and, consequently, increase the isoprenoid
precursor pool in E. coli. Carotenoids are a large class of biologically important compounds synthesized from isoprenoid precursors and of interest
for metabolic engineering. However, carotenoids are not ordinarily present in E. coli. Co-overexpression of E. coli dxs with Erwinia uredovora gene clusters encoding carotenoid biosynthetic enzymes led to an increased accumulation of the carotenoids lycopene or zeaxanthin
over controls not expressing DXS. Thus, rate-controlling enzymes encoded by the carotenogenic gene clusters are responsive
to an increase in isoprenoid precursor pools. Levels of accumulated carotenoids were increased up to 10.8 times the levels
of controls not overexpressing DXS. Lycopene accumulated to a level as high as 1333 μg/g dw and zeaxanthin accumulated to
a level as high as 592 μg/g dw, when pigments were extracted from colonies. Zeaxanthin-producing colonies grew about twice
as fast as lycopene-producing colonies throughout a time course of 11 days. Metabolic engineering of carbon flow from simple
glucose metabolites to representatives of the largest class of natural products was demonstrated in this model system.
Received: 6 August 1999 / Received revision: 25 October 1999 / Accepted: 5 November 1999 相似文献
7.
Albermann C 《Biotechnology letters》2011,33(2):313-319
The heterologous synthesis of lycopene in non-carotenogenic Escherichia coli required the introduction of the biosynthesis genes crtE, crtB, and crtI. Recombinant E. coli strains, expressing each lycopene biosynthesis gene from Pantoea ananatis using multi-copy plasmid or single-copies after stable chromosomal integration, were cultivated and the formation of lycopene
was investigated. The different expression conditions significantly influenced the lycopene formation as well as the growth
behaviour. High plasmid expression levels of crtI with a single copy background of crtE and crtB in E. coli led to a predominate synthesis of tetradehydrolycopene at 253 μg g−1 (cdw). 相似文献
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Tryptophan (Trp) halogenases are found in various bacteria and play an important role in natural product biosynthesis. Analysis
of the genome of Streptomyces toxytricini NRRL 15443 revealed an ORF, stth, encoding a putative Trp halogenase within a non-ribosomal peptide synthetase gene cluster. This gene was cloned into pET28a
and functionally overexpressed in Escherichia coli. The enzyme halogenated both l- and d-Trp to yield the corresponding 6-chlorinated derivatives. The optimum activity was at 40°C and pH 6 giving k
cat
/K
M value of STTH of 72,000 min−1 M−1. The enzyme also used bromide to yield 6-bromo-Trp. 相似文献
10.
Enrique A. Iturriaga Antonio Velayos Arturo P. Eslava 《Biotechnology and Bioprocess Engineering》2000,5(4):263-274
Carotenoids are widely distributed natural pigments which are in an increasing demand by the market, due to their applications
in the human food, animal feed, cosmetics, and pharmaceutical industries. Although more than 600 carotenoids have been identified
in nature, only a few are industrially important (β-carotene, astaxanthin, lutein or lycopene). To date chemical processes
manufacture most of the carotenoid production, but the interest for carotenoids of biological origin is growing since there
is an increased public concern over the safety of artificial food colorants. Although much interest and effort has been devoted
to the use of biological sources for industrially important carotenoids, only the production of biological β-carotene and
astaxanthin has been reported. Among fungi, several Mucorales strains, particularlyBlakeslea trispora, have been used to develop fermentation process for the production of β-carotene on almost competitive cost-price levels.
Similarly, the basidiomycetous yeastXanthophyllomyces dendrorhous (the perfect state ofPhaffia rhodozyma), has been proposed as a promising source of astaxanthin. This paper focuses on recent findings on the fungal pathways for
carotenoid production, especially the structure and function of the genes involved in the biosynthesis of carotenoids in the
Mucorales. An outlook of the possibilities of an increased industrial production of carotenoids, based on metabolic engineering
of fungi for carotenoid content and composition, is also discussed. 相似文献
11.
In the Tat protein export pathway of Gram-negative bacteria, TatA and TatB are homologous proteins that carry out distinct
and essential functions in separate sub-complexes. In contrast, Gram-positive Tat systems usually lack TatB and the TatA protein
is bifunctional. We have used a mutagenesis approach to delineate TatA/B-type domains in the bifunctional TatAd protein from
Bacillus subtilis. This involved expression of mutated TatAd variants in Escherichia coli and tests to determine whether the variants could function as TatA or TatB by complementing E. coli
tatA and/or tatB mutants. We show that mutations in the C-terminal half of the transmembrane span and the subsequent FGP ‘hinge’ motif are
critical for TatAd function with its partner TatCd subunit, and the same determinants are required for complementation of
either tatA or tatB mutants in Escherichia coli. This is thus a critical domain in both TatA and TatB proteins. In contrast, substitution of a series of residues at the
N-terminus specifically blocks the ability of TatAd to substitute for E. coli TatB. The results point to the presence of a universally conserved domain in the TatA/B-family, together with a separate
N-terminal domain that is linked to the TatB-type function in Gram-negative bacteria. 相似文献
12.
Rumila Mukherjee Shailen P. Borah Bhabesh C. Goswami 《Journal of applied phycology》2010,22(5):569-571
Two species of Trentepohlia, i.e., Trentepohlia aurea and Trentepohlia cucullata were collected from walls and tree bark, respectively, at two different seasons in a year. The total carotenoid content in
both the species is very high during winter but decreases significantly during summer. By spectroscopic analysis, it was found
that. T. aurea and T. cucullata growing in natural habitats are rich sources of carotenoids. The individual carotenoids were separated, identified, and estimated
by HPLC, and identified as β-carotene along with some other carotenoids, i.e., neoxanthin, lutein, β-cryptoxanthin, β,γ-carotene,
β,ε-carotene (absent during summer). 相似文献
13.
Boghigian BA Myint M Wu J Pfeifer BA 《Journal of industrial microbiology & biotechnology》2011,38(11):1809-1820
Natural products have long served as rich sources of drugs possessing a wide range of pharmacological activities. The discovery
and development of natural product drug candidates is often hampered by the inability to efficiently scale and produce a molecule
of interest, due to inherent qualities of the native producer. Heterologous biosynthesis in an engineering and process-friendly
host emerged as an option to produce complex natural products. Escherichia coli has previously been utilized to produce complex precursors to two popular natural product drugs, erythromycin and paclitaxel.
These two molecules represent two of the largest classes of natural products, polyketides and isoprenoids, respectively. In
this study, we have developed a platform E. coli strain capable of simultaneous production of both product precursors at titers greater than 15 mg l−1. The utilization of a two-phase batch bioreactor allowed for very strong in situ separation (having a partitioning coefficient
of greater than 5,000), which would facilitate downstream purification processes. The system developed here could also be
used in metagenomic studies to screen environmental DNA for natural product discovery and preliminary production experiments. 相似文献
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Cyanobacteria produce some carotenoids. We identified the molecular structures, including the stereochemistry, of all the
carotenoids in the terrestrial cyanobacterium, Nostoc commune NIES-24 (IAM M-13). The major carotenoid was β-carotene. Its hydroxyl derivatives were (3R)-β-cryptoxanthin, (3R,3′R)-zeaxanthin, (2R,3R,3′R)-caloxanthin, and (2R,3R,2′R,3′R)-nostoxanthin, and its keto derivatives were echinenone and canthaxanthin. The unique myxol glycosides were (3R,2′S)-myxol 2′-fucoside and (2R,3R,2′S)-2-hydroxymyxol 2′-fucoside. This is only the second species found to contain 2-hydroxymyxol. We propose possible carotenogenesis
pathways based on our identification of the carotenoids: the hydroxyl pathway produced nostoxanthin via zeaxanthin from β-carotene,
the keto pathway produced canthaxanthin from β-carotene, and the myxol pathway produced 2-hydroxymyxol 2′-fucoside via myxol
2′-fucoside. This cyanobacterium was found to contain many kinds of carotenoids and also displayed many carotenogenesis pathways,
while other cyanobacteria lack some carotenoids and a part of carotenogenesis pathways compared with this cyanobacterium. 相似文献
16.
Harada H Yu F Okamoto S Kuzuyama T Utsumi R Misawa N 《Applied microbiology and biotechnology》2009,81(5):915-925
We show here an efficient synthesis system of isoprenoids from acetoacetate as the main substrate. We expressed in Escherichia coli a Streptomyces mevalonate pathway gene cluster starting from HMG-CoA synthase and including isopentenyl diphosphate isomerase (idi) type 2 gene and the yeast idi type 1 and rat acetoacetate-CoA ligase (Aacl) genes. When the α-humulene synthase (ZSS1) gene of shampoo ginger was expressed in this transformant, the resultant E. coli produced 958 μg/mL culture of α-humulene with a lithium acetoacetate (LAA) supplement, which was a 13.6-fold increase compared
with a control E. coli strain expressing only ZSS1. Next, we investigated if this E. coli strain engineered to utilize acetoacetate can synthesize carotenoids effectively. When the crtE, crtB, and crtI genes required for lycopene synthesis were expressed in the transformant, lycopene amounts reached 12.5 mg/g dry cell weight
with addition of LAA, an 11.8-fold increase compared with a control expressing only the three crt genes. As for astaxanthin production with the E. coli transformant, in which the crtE, crtB, crtI, crtY, crtZ, and crtW genes were expressed, the total amount of carotenoids produced (astaxanthin, lycopene, and phytoene) was significantly increased
to 7.5 times that of a control expressing only the six crt genes. 相似文献
17.
Flower color alteration in <Emphasis Type="Italic">Lotus japonicus</Emphasis> by modification of the carotenoid biosynthetic pathway 总被引:1,自引:0,他引:1
Suzuki S Nishihara M Nakatsuka T Misawa N Ogiwara I Yamamura S 《Plant cell reports》2007,26(7):951-959
To establish a model system for alteration of flower color by carotenoid pigments, we modified the carotenoid biosynthesis
pathway of Lotus japonicus using overexpression of the crtW gene isolated from marine bacteria Agrobacterium aurantiacum and encoding β-carotene ketolase (4,4′-β-oxygenase) for the production of pink to red color ketocarotenoids. The crtW gene with the transit peptide sequence of the pea Rubisco small subunit under the regulation of the CaMV35S promoter was
introduced to L. japonicus. In most of the resulting transgenic plants, the color of flower petals changed from original light yellow to deep yellow
or orange while otherwise exhibiting normal phenotype. HPLC and TLC analyses revealed that leaves and flower petals of these
plants accumulated novel carotenoids, believed to be ketocarotenoids consisting of including astaxanthin, adonixanthin, canthaxanthin
and echinenone. Results indicated that modification of the carotenoid biosynthesis pathway is a means of altering flower color
in ornamental crops. 相似文献
18.
Kaneko M Hwang EI Ohnishi Y Horinouchi S 《Journal of industrial microbiology & biotechnology》2003,30(8):456-461
Chalcones, the central precursor of flavonoids, are synthesized exclusively in plants from tyrosine and phenylalanine via
the sequential reaction of phenylalanine ammonia-lyase (PAL), cinnamate-4-hydroxylase (C4H), 4-coumarate:coenzyme A ligase
(4CL) and chalcone synthase (CHS). Chalcones are converted into the corresponding flavanones by the action of chalcone isomerase
(CHI), or non-enzymatically under alkaline conditions. PAL from the yeast Rhodotorula rubra, 4CL from an actinomycete Streptomyces coelicolor A3(2), and CHS from a licorice plant Glycyrrhiza echinata, assembled as artificial gene clusters in different organizations, were used for fermentation production of flavanones in Escherichia coli. Because the bacterial 4CL enzyme attaches CoA to both cinnamic acid and 4-coumaric acid, the designed biosynthetic pathway
bypassed the C4H step. E. coli carrying one of the designed gene clusters produced about 450 μg naringenin/l from tyrosine and 750 μg pinocembrin/l from
phenylalanine. The successful production of plant-specific flavanones in bacteria demonstrates the usefulness of combinatorial
biosynthesis approaches not only for the production of various compounds of plant and animal origin but also for the construction
of libraries of "unnatural" natural compounds.
Dedicated to Professor Sir David Hopwood. 相似文献
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Neomycin and paromomycin are aminoglycoside antibiotics that specifically stimulate the misreading of mRNA by binding to the
decoding site of 16S rRNA in the 30S ribosomal subunit. Recent work has shown that both antibiotics also inhibit 30S subunit
assembly in Escherichia coli and Staphylococcus aureus cells. This work describes the characteristics of an assembly intermediate produced in E. coli cells grown with neomycin or paromomycin. Antibiotic treatment stimulated the accumulation of a 30S assembly precursor with
a sedimentation coefficient of 21S. The particle was able to bind radio-labeled antibiotics in vivo and in vitro. Hybridization
experiments showed that the 21S precursor particle contained unprocessed 16S rRNA with both 5′ and 3′ extensions. Ten 30S
ribosomal proteins were found in the precursor after inhibition by each drug. In addition, cell free reconstitution assays
generated a 21S particle after incubation with either aminoglycoside. This work helps to define the features of the ribosome
structure as a target for antimicrobial agents and may provide information needed for the design of more effective antibiotics. 相似文献