Abstract:
:Pathway refactoring serves as an invaluable synthetic biology tool for natural product discovery, characterization, and engineering. However, the complicated and laborious molecular biology techniques largely hinder its application in natural product research, especially in a high-throughput manner. Here we report a plug-and-play pathway refactoring workflow for high-throughput, flexible pathway construction, and expression in both Escherichia coli and Saccharomyces cerevisiae. Biosynthetic genes were firstly cloned into pre-assembled helper plasmids with promoters and terminators, resulting in a series of expression cassettes. These expression cassettes were further assembled using Golden Gate reaction to generate fully refactored pathways. The inclusion of spacer plasmids in this system would not only increase the flexibility for refactoring pathways with different number of genes, but also facilitate gene deletion and replacement. As proof of concept, a total of 96 pathways for combinatorial carotenoid biosynthesis were built successfully. This workflow should be generally applicable to different classes of natural products produced by various organisms. Biotechnol. Bioeng. 2017;114: 1847-1854. © 2017 Wiley Periodicals, Inc.
journal_name
Biotechnol Bioengjournal_title
Biotechnology and bioengineeringauthors
Ren H,Hu P,Zhao Hdoi
10.1002/bit.26309subject
Has Abstractpub_date
2017-08-01 00:00:00pages
1847-1854issue
8eissn
0006-3592issn
1097-0290journal_volume
114pub_type
杂志文章abstract::Purified rat liver phenylalanine hydroxylase [L-phenylalanine:tetrahydropteridine:oxygen oxidoreductase (4-hydroxylating), EC 1.14.16.1] was immobilized with activated thiol-Sepharose 4B via disulfide bond formation, which is expected to immobilize the enzyme in its activated form through the SH modification. This imm...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260280422
更新日期:1986-04-01 00:00:00
abstract::High-level expression of recombinant penicillin acylase (PAC) using the strong trc promoter system in Escherichia coli is frequently limited by the processing and folding of PAC precursors (proPAC) in the periplasm, resulting in physiological stress and inclusion body formation in this compartment. Periplasmic heat-sh...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.21161
更新日期:2007-04-01 00:00:00
abstract::Product inhibition of biological production systems is a widely observed phenomenon with prominent implications for the design and ultimately the success of biotechnological processes. In order to investigate whether such effects could limit the maximal concentrations in the production of the beta-lactamase inhibitor ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.21783
更新日期:2008-06-15 00:00:00
abstract::This study describes the development and application of a bioaffinity chromatographic system for the one-step purification of an NADP(+)-dependent secondary alcohol dehydrogenase from the obligate anaerobe, Thermoanaerobacter brockii (TBADH, EC 1.1.1.2). The general approach is based upon improving the selectivity of ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.10079
更新日期:2002-03-05 00:00:00
abstract::Ethanol-producing bioreactors employing cells of Zymomonas mobilis attached to glass-fiber pads were operated continuously for as long as 28 days. Ethanol production, which is related to bed-associated biomass levels, was found to occur in three distinct phases: an exponential phase, a linear phase, and a "steady-stat...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260240306
更新日期:1982-03-01 00:00:00
abstract::Recent studies have demonstrated the feasibility of using ultrafiltration for the purification of plasmid DNA, but there is still little understanding of the factors governing DNA transmission. Experimental data were obtained for the transmission of a 3.0 kbp supercoiled plasmid DNA through composite regenerated cellu...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.21575
更新日期:2008-02-01 00:00:00
abstract::A pure culture of Pseudomonas fluorescens was used as a model system to study the kinetics of denitrification. An exponentially growing culture was harvested and resuspended in an anoxic acetate solution buffered with K/Na phosphate at pH values of 6.6, 7.0, 7.4, and 7.8. The temperature was kept at 28 degrees C in al...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260460512
更新日期:1995-06-05 00:00:00
abstract::Tissue engineering devices as in vitro cell culture systems in scaffolds has encountered the bottleneck due to their much lower cell functions than real tissues/organs in vivo. Such situation has been improved in some extent by mimicking the cell microenvironments in vivo from either chemical or physical ways. However...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.24874
更新日期:2013-08-01 00:00:00
abstract::The release kinetics of beta-galactosidase protein have been determined during small-scale ultrasonication of E. coli cells. Among several studied parameters, ionic strength and cell concentration have the least influence on the rate of protein recovery, whereas sample volume and acoustic power dramatically affect the...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/(sici)1097-0290(19980605)58:5<536::aid-bit
更新日期:1998-06-05 00:00:00
abstract::We propose that transcript levels for some genes are affected by the bacterial cell division cycle and this may be an important factor to consider when designing synthetic bacterial genomes. To test this hypothesis, transcript levels of 58 genes in Escherichia coli B/r A were determined at five times during the cell d...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.22098
更新日期:2009-02-15 00:00:00
abstract::Anaerobic biological wastewater treatment has numerous advantages over conventional aerobic processes; anaerobic biotechnologies, however, still have a reputation for low-quality effluents and operational instabilities. In this study, anaerobic bioreactors were augmented with an oxygen-transferring membrane to improve...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.20219
更新日期:2005-02-20 00:00:00
abstract::Extracellular beta-xylosidase (1,4-beta-D-xylan xylohydrolase, EC 3.2.1.37) from culture filtrates of Neurospora crassa was purified to homogeneity by preparative isoelectric focusing followed by gel electrophoresis. The molecular weight of the purified xylosidase was 83,000 D and the K(m) on p-nitrophenyl-beta-D-xylo...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260281210
更新日期:1986-12-01 00:00:00
abstract::The sufficient provision of oxygen is mandatory for enzymatic oxidations in aqueous solution, however, in process optimization this still is a bottleneck that cannot be overcome with the established methods of macrobubble aeration. Providing higher mass transfer performance through microbubble aerators, inefficient ae...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.27556
更新日期:2021-01-01 00:00:00
abstract::Aspergillus terreus cadA, encoding cis-aconitate decarboxylase, is an essential gene for itaconic acid (IA) biosynthesis, but it is primarily expressed as insoluble aggregates in most industrial hosts. This has been a hurdle for the development of recombinant strategies for IA production. Here, we created a library of...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.25914
更新日期:2016-07-01 00:00:00
abstract::Wastewater-fed microbial fuel cells (MFCs) are a promising technology to treat low-organic carbon wastewater and recover part of the chemical energy in wastewater as electrical power. However, the interactions between electrochemically active and fermentative microorganisms cannot be easily studied in wastewater-fed M...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.25137
更新日期:2014-04-01 00:00:00
abstract::Light-responsive bioconjugates are synthesized by a two-step protocol calling first for cotelomerization (chain-transfer polymerization) of N-isopropylacrylamide and N-acryloxysuccinimide. The desired bioligand (biotin) is used in modified form as chain-transfer agent in this step. As a consequence, 100% of the produc...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.20479
更新日期:2005-09-05 00:00:00
abstract::By feeding ethanol at various high rates to low cell density cultures of Saccharomyces cerevisiae it was shown that the sharp fall in viability when ethanol is produced during rapid fermentations is in part a direct consequence of the high rate of change of extracellular ethanol concentration. Nevertheless, the fall i...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260350202
更新日期:1990-01-20 00:00:00
abstract::Hyperacute rejection following xenogeneic transplantation in primates is mediated by naturally occurring IgM antibodies, which are specifically directed to alpha-Galactosyl residues on many nonprimate mammalian cells. Current approaches to remove these anti-alphaGal IgM include plasmapheresis followed by immunoaffinit...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.10555
更新日期:2003-01-20 00:00:00
abstract::We demonstrate a novel application of (13)C pulsed field gradient (PFG) NMR to monitor mass transfer, due to both flow and diffusion, in a 3D complex porous support structure modified by biofilm growth. This enables timescales an order of magnitude larger than previously possible to be accessed with respect to displac...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.21913
更新日期:2008-10-15 00:00:00
abstract::The biological molecule responsible for the suppression of pyritic sulfur in fine coal simulated froth flotation treated with bacteria was identified. Protein was found to be the most effective agent in pyrite suppression of the three cell components (protein, lipid, and carbohydrate) assayed. Coal recovery and ash re...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260330606
更新日期:1989-02-05 00:00:00
abstract::Metabolic engineering of isopropyl alcohol (IPA)-producing Escherichia coli strains was conducted along with 13 C-metabolic flux analysis (MFA). A metabolically engineered E. coli strain expressing the adc gene derived from Clostridium acetobutylicum and the IPADH gene from C. beijerinckii did not produce IPA during i...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.26390
更新日期:2017-12-01 00:00:00
abstract::Being biosurfactants, rhamnolipids create severe foaming when produced in aerobic Pseudomonas aeruginosa fermentation. The necessary reduction of aeration causes oxygen limitation and restricts cell and product concentrations. In this study, we evaluate the new strategy of rhamnolipid production under denitrification ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/1097-0290(20010105)72:1<25::aid-bit4>3.0.c
更新日期:2001-01-05 00:00:00
abstract::Pre-filtration using ion exchange membrane adsorbers can improve parvovirus filter throughput of monoclonal antibodies (mAbs). The membranes work by binding trace foulants, and although some antibody product also binds, yields > or =99% are easily achieved by overloading. Results show that foulant adsorption is depend...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.22729
更新日期:2010-07-01 00:00:00
abstract::Reactive oxygen species (ROS), encompassing all oxygen radical or non-radical oxidizing agents, play key roles in disease progression. Controlled delivery of antioxidants is therapeutically relevant in such oxidant-stressed environments. Encapsulating small hydrophilic molecules into hydrophobic polymer microparticles...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.26443
更新日期:2018-01-01 00:00:00
abstract::Alginate gels produced by an external or internal gelation technique were studied so as to determine the optimal bead matrix within which DNA can be immobilized for in vivo application. Alginates were characterized for guluronic/mannuronic acid (G/M) content and average molecular weight using 1H-NMR and LALLS analysis...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:
更新日期:1998-02-20 00:00:00
abstract::We report the isolation of mutant strains of the methylotrophic yeast Hansenula polymorpha that are able to efficiently oxidize ethanol to acetaldehyde in an intact cell system. The oxidation reaction is catalyzed by alcohol oxidase (AOX), a key enzyme in the methanol metabolic pathway that is typically present only i...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/(sici)1097-0290(20000405)68:1<44::aid-bit5
更新日期:2000-04-05 00:00:00
abstract::A mathematical model is formulated to describe trends in biomass and penicillin formation as well as substrate consumption for fed-batch cultivations. The biomass is structured into three morphological compartments, and glucose and corn steep liquor are considered as substrates for growth. Penicillin formation is assu...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/(SICI)1097-0290(19971220)56:6<593::AID-BIT
更新日期:1997-12-20 00:00:00
abstract::An immobilized enzyme reactor has been developed to remove heparin, the anticoagulant that is required in all extracorporeal devices for patients undergoing open-heart surgery or kidney dialysis. The device uses the enzyme heparinase (EC 4.2.2.7), which is covalently linked to agarose with cyanogen bromide. A critical...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260300209
更新日期:1987-08-05 00:00:00
abstract::CO(2) partial pressure (pCO(2)) in industrial cell culture reactors may reach 150-200 mm Hg, which can significantly inhibit cell growth and recombinant protein production. The inhibitory effects of elevated pCO(2) at constant pH are due to a combination of the increases in pCO(2) and [HCO(-) (3)], per se, and the ass...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.10176
更新日期:2002-02-15 00:00:00
abstract::Somatic embryogenesis is among the most promising means of large scale plant micropropagation. The development of somatic embryos is characterized by their morphological changes. Embryos in culture usually exhibit high heterogeneity and abnormality. As conventional microscopic observation is laborious and subjective, ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/(SICI)1097-0290(19960405)50:1<65::AID-BIT8
更新日期:1996-04-05 00:00:00