Bioengineering of oleaginous yeast Yarrowia lipolytica for lycopene production.

Oleaginous yeast Yarrowia lipolytica is capable of accumulating large amount of lipids. There is a growing interest to engineer this organism to produce lipid-derived compounds for a variety of applications. In addition, biosynthesis of value-added products such as carotenoid and its derivatives have been explored.

Construction of carotenoid biosynthetic pathways through chromosomal integration in methane-utilizing bacterium Methylomonas sp. strain 16a.

Methylomonas sp. strain 16a is an obligate methanotrophic bacterium that uses methane or methanol as the sole energy and carbon source. In order to engineer a stable strain to produce carotenoids, integration of genes or gene clusters in various nonessential locations in the chromosome is used.

Use of transposon promoter-probe vectors in the metabolic engineering of the obligate methanotroph Methylomonas sp. strain 16a for enhanced C40 carotenoid synthesis.

The recent expansion of genetic and genomic tools for metabolic engineering has accelerated the development of microorganisms for the industrial production of desired compounds. We have used transposable elements to identify chromosomal locations in the obligate methanotroph Methylomonas sp. strain 16a that support high-level expression of genes involved in the synthesis of the C(40) carotenoids canthaxanthin and astaxanthin.

Construction of the astaxanthin biosynthetic pathway in a methanotrophic bacterium Methylomonas sp. strain 16a.

Methylomonas sp. strain 16a is an obligate methanotrophic bacterium that uses methane or methanol as the sole carbon source. An effort was made to engineer this organism for astaxanthin production.

Expression of bacterial hemoglobin genes to improve astaxanthin production in a methanotrophic bacterium Methylomonas sp.

Astaxanthin has been widely used as a feed supplement in poultry and aquaculture industries. One challenge for astaxanthin production in bacteria is the low percentage of astaxanthin in the total carotenoids. An obligate methanotrophic bacterium Methylomonas sp.

Mutational and functional analysis of the beta-carotene ketolase involved in the production of canthaxanthin and astaxanthin.

Biosynthesis of the commercial carotenoids canthaxanthin and astaxanthin requires beta-carotene ketolase. The functional importance of the conserved amino acid residues of this enzyme from Paracoccus sp. strain N81106 (formerly classified as Agrobacterium aurantiacum) was analyzed by alanine-scanning mutagenesis.

Genetic and physiological responses of Bacillus subtilis to metal ion stress.

Metal ion homeostasis is regulated principally by metalloregulatory proteins that control metal ion uptake, storage and efflux genes. We have used transcriptional profiling to survey Bacillus subtilis for genes that are rapidly induced by exposure to high levels of metal ions including Ag(I), Cd(II), Cu(II), Ni(II) and Zn(II) and the metalloid As(V). Many of the genes affected by metal stress were controlled by known metalloregulatory proteins (Fur, MntR, PerR, ArsR and CueR).

Differential gene expression shows natural brominated furanones interfere with the autoinducer-2 bacterial signaling system of Escherichia coli.

The quorum sensing disrupter (5Z)-4-bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone (furanone) of the alga Delisea pulchra was previously found by us (Environ Microbiol 3:731-736, 2001) to inhibit quorum sensing in Escherichia coli via autoinducer-2 (AI-2, produced by LuxS). In this study, DNA microarrays were used to study the genetic basis of this natural furanone inhibition of AI-2 signaling (significant values with p < 0.05 are reported).

Bacillus subtilis ResD induces expression of the potential regulatory genes yclJK upon oxygen limitation.

Transcription of the yclJK operon, which encodes a potential two-component regulatory system, is activated in response to oxygen limitation in Bacillus subtilis. Northern blot analysis and assays of yclJ-lacZ reporter gene fusion activity revealed that the anaerobic induction is dependent on another two-component signal transduction system encoded by resDE. ResDE was previously shown to be required for the induction of anaerobic energy metabolism.

Differential gene expression to investigate the effect of (5Z)-4-bromo- 5-(bromomethylene)-3-butyl-2(5H)-furanone on Bacillus subtilis.

(5Z)-4-Bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone (furanone) from the red marine alga Delisea pulchra was found previously to inhibit the growth, swarming, and biofilm formation of gram-positive bacteria. Using the gram-positive bacterium Bacillus subtilis as a test organism, we observed cell killing by 20 microg of furanone per ml, while 5 microg of furanone per ml inhibited growth approximately twofold without killing the cells. To discover the mechanism of this inhibition on a genetic level and to investigate furanone as a novel antibiotic, full-genome DNA microarrays were used to analyze the gene expression profiles of B.

Response of Bacillus subtilis to nitric oxide and the nitrosating agent sodium nitroprusside.

We examined the effects of nitric oxide (NO) and sodium nitroprusside (SNP) on Bacillus subtilis physiology and gene expression. In aerobically growing cultures, cell death was most pronounced when NO gas was added incrementally rather than as a single bolus, suggesting that the length of exposure was important in determining cell survival. DNA microarrays, Northern hybridizations, and RNA slot blot analyses were employed to characterize the global transcriptional response of B.

Gene expression in Bacillus subtilis surface biofilms with and without sporulation and the importance of yveR for biofilm maintenance.

Five independent DNA microarray experiments were used to study the gene expression profile of a 5-day Bacillus subtilis air-liquid interface biofilm relative to planktonic cells. Both wild-type B. subtilis and its sporulation mutant (DeltaspoIIGB::erm) were investigated to discern the important biofilm genes (in the presence and absence of sporulation).

Stationary-phase quorum-sensing signals affect autoinducer-2 and gene expression in Escherichia coli.

Quorum sensing via autoinducer-2 (AI-2) has been identified in different strains, including those from Escherichia, Vibrio, Streptococcus, and Bacillus species, and previous studies have suggested the existence of additional quorum-sensing signals working in the stationary phase of Escherichia coli cultures. To investigate the presence and global effect of these possible quorum-sensing signals other than AI-2, DNA microarrays were used to study the effect of stationary-phase signals on the gene expression of early exponential-phase cells of the AI-2-deficient strain E. coli DH5alpha.

Cell wall stress responses in Bacillus subtilis: the regulatory network of the bacitracin stimulon.

In response to sublethal concentrations of antibiotics, bacteria often induce an adaptive response that can contribute to antibiotic resistance. We report the response of Bacillus subtilis to bacitracin, an inhibitor of cell wall biosynthesis found in its natural environment. Analysis of the global transcriptional profile of bacitracin-treated cells reveals a response orchestrated by two alternative sigma factors (sigmaB and sigmaM) and three two-component systems (YvqEC, YvcPQ and BceRS).

The global transcriptional response of Bacillus subtilis to manganese involves the MntR, Fur, TnrA and sigmaB regulons.

We have used DNA microarrays to monitor the global transcriptional response of Bacillus subtilis to changes in manganese availability. Mn(II) leads to the MntR-dependent repression of both the mntH and mntABCD operons encoding Mn(II) uptake systems. Mn(II) also represses the Fur regulon.

Regulation of the Bacillus subtilis extracytoplasmic function protein sigma(Y) and its target promoters.

The Bacillus subtilis extracytoplasmic function sigma factor sigma(Y) is of unknown function. We demonstrate that the sigY operon is expressed from an autoregulatory promoter site, P(Y). We selected for transposon-induced mutations that upregulate P(Y) transcription in an attempt to identify genes involved in sigma(Y) regulation.

Functional analysis of the Bacillus subtilis Zur regulon.

The Bacillus subtilis zinc uptake repressor (Zur) regulates genes involved in zinc uptake. We have used DNA microarrays to identify genes that are derepressed in a zur mutant. In addition to members of the two previously identified Zur-regulated operons (yciC and ycdHI-yceA), we identified two other genes, yciA and yciB, as targets of Zur regulation.