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Ingemar N?rdal Johannes Pfeifenschneider Trygve Brautaset Volker F Wendisch 《Microbial biotechnology》2015,8(2):342-350
Methanol is regarded as an attractive substrate for biotechnological production of value-added bulk products, such as amino acids and polyamines. In the present study, the methylotrophic and thermophilic bacterium Bacillus methanolicus was engineered into a microbial cell factory for the production of the platform chemical 1,5-diaminopentane (cadaverine) from methanol. This was achieved by the heterologous expression of the Escherichia coli genes cadA and ldcC encoding two different lysine decarboxylase enzymes, and by increasing the overall L-lysine production levels in this host. Both CadA and LdcC were functional in B. methanolicus cultivated at 50°C and expression of cadA resulted in cadaverine production levels up to 500 mg l−1 during shake flask conditions. A volume-corrected concentration of 11.3 g l−1 of cadaverine was obtained by high-cell density fed-batch methanol fermentation. Our results demonstrated that efficient conversion of L-lysine into cadaverine presumably has severe effects on feedback regulation of the L-lysine biosynthetic pathway in B. methanolicus. By also investigating the cadaverine tolerance level, B. methanolicus proved to be an exciting alternative host and comparable to the well-known bacterial hosts E. coli and Corynebacterium glutamicum. This study represents the first demonstration of microbial production of cadaverine from methanol. 相似文献
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Steffen N. Lindner Sandra Knebel Srinivas R. Pallerla Siegfried M. Schoberth Volker F. Wendisch 《Applied microbiology and biotechnology》2010,87(2):703-713
The Corynebacterium glutamicum gene cg2091 is encoding a polyphosphate (PolyP)/ATP-dependent glucokinase (PPGK). Previous work demonstrated the association
of PPGK to PolyP granules. The deduced amino acid sequence of PPGK shows 45% sequence identity to PolyP/ATP glucomannokinase
of Arthrobacter sp. strain KM and 50% sequence identity to PolyP glucokinase of Mycobacterium tuberculosis H37Rv. PPGK from C. glutamicum was purified from recombinant Escherichia coli. PolyP was highly preferred over ATP and other NTPs as substrate and with respect to the tested PolyPs differing in chain
length; the protein was most active with PolyP75. Gel filtration analysis revealed that PolyP supported the formation of homodimers of PPGK and that PPGK was active as a
homodimer. A ppgK deletion mutant (ΔppgK) showed slowed growth in minimal medium with maltose as sole carbon source. Moreover, in minimal medium containing 2 to 4%
(w/v) glucose as carbon source, ΔppgK grew to lower final biomass concentrations than the wild type. Under phosphate starvation conditions, growth of ΔppgK was reduced, and growth of a ppgK overexpressing strain was increased as compared to wild type and empty vector control, respectively. Thus, under conditions
of glucose excess, the presence of PPGK entailed a growth advantage. 相似文献
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Osamu Kato Jung-Won Youn K Corinna Stansen Daisuke Matsui Tadao Oikawa Volker F Wendisch 《BMC microbiology》2010,10(1):321
Background
Corynebacterium glutamicum is able to grow with lactate as sole or combined carbon and energy source. Quinone-dependent L-lactate dehydrogenase LldD is known to be essential for utilization of L-lactate by C. glutamicum. D-lactate also serves as sole carbon source for C. glutamicum ATCC 13032. 相似文献6.
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We investigated the global gene expression changes of Escherichia coli due to the presence of different concentrations of phenylalanine or shikimate in the growth medium. The response to 0.5 g l(-1) phenylalanine primarily reflected a perturbed aromatic amino acid metabolism, in particular due to TyrR-mediated regulation. The addition of 5g l(-1) phenylalanine reduced the growth rate by half and elicited a great number of likely indirect effects on genes regulated in response to changed pH, nitrogen or carbon availability. Consistent with the observed gene expression changes, supplementation with shikimate, tyrosine and tryptophan relieved growth inhibition by phenylalanine. In contrast to the wild-type, a tyrR disruption strain showed increased expression of pckA and of tktB in the presence of phenylalanine, but its growth was not affected by phenylalanine at the concentrations tested. The absence of growth inhibition by phenylalanine suggested that at high phenylalanine concentrations TyrR-defective strains might perform better in phenylalanine production. 相似文献
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Characterization of myo-inositol utilization by Corynebacterium glutamicum: the stimulon, identification of transporters, and influence on L-lysine formation
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Krings E Krumbach K Bathe B Kelle R Wendisch VF Sahm H Eggeling L 《Journal of bacteriology》2006,188(23):8054-8061
Although numerous bacteria possess genes annotated iol in their genomes, there have been very few studies on the possibly associated myo-inositol metabolism and its significance for the cell. We found that Corynebacterium glutamicum utilizes myo-inositol as a carbon and energy source, enabling proliferation with a high maximum rate of 0.35 h-1. Whole-genome DNA microarray analysis revealed that 31 genes respond to myo-inositol utilization, with 21 of them being localized in two clusters of >14 kb. A set of genomic mutations and functional studies yielded the result that some genes in the two clusters are redundant, and only cluster I is necessary for catabolizing the polyol. There are three genes which encode carriers belonging to the major facilitator superfamily and which exhibit a >12-fold increased mRNA level on myo-inositol. As revealed by mutant characterizations, one carrier is not involved in myo-inositol uptake whereas the other two are active and can completely replace each other with apparent Kms for myo-inositol as a substrate of 0.20 mM and 0.45 mM, respectively. Interestingly, upon utilization of myo-inositol, the L-lysine yield is 0.10 mol/mol, as opposed to 0.30 mol/mol, with glucose as the substrate. This is probably not only due to myo-inositol metabolism alone since a mixture of 187 mM glucose and 17 mM myo-inositol, where the polyol only contributes 8% of the total carbon, reduced the L-lysine yield by 29%. Moreover, genome comparisons with other bacteria highlight the core genes required for growth on myo-inositol, whose metabolism is still weakly defined. 相似文献
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Thermoproteus tenax is a hyperthermophilic, facultative heterotrophic archaeum. In this organism the utilization of the two catabolic pathways,
a variant of the Embden-Meyerhof-Parnas (EMP) pathway and the modified (nonphosphorylative) Entner-Doudoroff (ED) pathway,
was investigated and the first enzyme of the ED pathway, glucose dehydrogenase, was characterized. The distribution of the
13C label in alanine synthesized by cells grown with [1-13C]glucose indicated that in vivo the EMP pathway and the modified ED pathway operate parallel, with glucose metabolization
via the EMP pathway being prominent. To initiate studies on the regulatory mechanisms governing carbon flux via these pathways,
the first enzyme of the ED pathway, glucose dehydrogenase, was purified to homogeneity and its phenotypic properties were
characterized. The pyridine-nucleotide-dependent enzyme used both NAD+ and NADP+ as cosubstrates, showing a 100-fold higher affinity for NADP+. Besides glucose, xylose was used as substrate, but with significantly lower affinity. These data suggest that the physiological
function of the enzyme is the oxidation of glucose by NADP+. A striking feature was the influence of NADP+ and NAD+ on the quaternary structure and activity state of the enzyme. Without cosubstrate, the enzyme was highly aggregated (mol.
mass > 600 kDa) but inactive, whereas in the presence of the cosubstrate the aggregates dissociated into enzymatically active,
homomeric dimers with a mol. mass of 84 kDa (mol. mass of subunits: 41 kDa). The N-terminal amino acid sequence showed striking
similarity to the respective partial sequences of alcohol dehydrogenases and sorbitol dehydrogenases, but no resemblance to
the known pyridine-nucleotide-dependent archaeal and bacterial glucose dehydrogenases.
Received: 25 October 1996 / Accepted: 15 April 1997 相似文献
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Characterization of a Corynebacterium glutamicum Lactate Utilization Operon Induced during Temperature-Triggered Glutamate Production 总被引:1,自引:0,他引:1
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