A Colorimetric Determination of Blood Acetoin

Westerfeld, W. W.

doi:10.1016/s0021-9258(17)41484-0

Cited by 638 publications

(38 citation statements)

References 16 publications

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“…The mixture was incubated for an additional 25 min at 37°C to allow for the acid hydrolysis of the acetolactate to acetoin. The acetoin formed was then measured using the Voges-Proskauer test (Eddy, 1961;Westerfeld, 1945) A 0.2-mL aliquot of the aforementioned mixture was transferred to a tube with 0.8 mL of 0.45N NaOH. A 1-mL aliquot of 1:1 volume ratio ␣-naphthol (5% ␣-naphthol in 2.5N NaOH) to 0.5% creatine was added and the resulting mixture maintained at room temperature for 30 min with shaking.…”

Section: B Subtilis Als Assaymentioning

confidence: 99%

Effect of variation ofKlebsiella pneumoniae acetolactate synthase expression on metabolic flux redistribution inEscherichia coli

Yang

Peredelchuk

Bennett

et al. 2000

Biotechnol. Bioeng.

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Escherichia coli strains carrying the Bacillus subtilis acetolactate synthase (ALS) gene were previously shown to produce less acetate with higher ATP yields. Metabolic flux analysis was used to show that excess pyruvate was channeled into the less inhibitory product, acetoin. To further understand the role of intrinsic enzymatic properties and the effect of variations in enzyme levels in the alternation of metabolic fluxes, we constructed a chromosomal integrant of the Klebsiella pneumoniae ALS gene. The reported in vitro Michaelis-Menten constants (K m ) for the Bacillus and the Klebsiella ALS are 13.0 mM and 8.0 mM, respectively. Furthermore, expression of the Klebsiella ALS is under the control of an inducible trp promoter system. Shake-flask experiments showed a linear induction response (the ALS activity changes from about 9 to 223 U/mg of protein when the inducer concentration [IAA] varied from 0 to 40 mg/ L). Chemostat experiments showed a similar induction response. Interactions between the branched reactions catalyzed by the PFL, LDH, and the ALS enzymes at the pyruvate node were examined. The results indicate the importance of in vivo enzyme activities in the redistribution of metabolic fluxes.

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Section: B Subtilis Als Assaymentioning

confidence: 99%

Effect of variation ofKlebsiella pneumoniae acetolactate synthase expression on metabolic flux redistribution inEscherichia coli

Yang

Peredelchuk

Bennett

et al. 2000

Biotechnol. Bioeng.

View full text Add to dashboard Cite

show abstract

“…Acetolactate synthase activity was tested as described previously [ 25 ]. Acetolactate produced after one hour was assayed at a single endpoint by conversion to acetoin, which was detected using the reaction method described by Westerfeldt [ 26 ] and quantified through the use of a standard curve after subtraction of the acetoin produced without substrate. The acetoin content was then normalized based on the protein content.…”

Section: Methodsmentioning

confidence: 99%

Effects of Mixing Volatile Fatty Acids as Carbon Sources on Rhodospirillum rubrum Carbon Metabolism and Redox Balance Mechanisms

et al. 2021

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Rhodospirillum rubrum has a versatile metabolism, and as such can assimilate a broad range of carbon sources, including volatile fatty acids. These carbon sources are gaining increasing interest for biotechnological processes, since they reduce the production costs for numerous value-added compounds and contribute to the development of a more circular economy. Usually, studies characterizing carbon metabolism are performed by supplying a single carbon source; however, in both environmental and engineered conditions, cells would rather grow on mixtures of volatile fatty acids (VFAs) generated via anaerobic fermentation. In this study, we show that the use of a mixture of VFAs as carbon source appears to have a synergy effect on growth phenotype. In addition, while propionate and butyrate assimilation in Rs. rubrum is known to require an excess of bicarbonate in the culture medium, mixing them reduces the requirement for bicarbonate supplementation. The fixation of CO2 is one of the main electron sinks in purple bacteria; therefore, this observation suggests an adaptation of both metabolic pathways used for the assimilation of these VFAs and redox homeostasis mechanism. Based on proteomic data, modification of the propionate assimilation pathway seems to occur with a switch from a methylmalonyl-CoA intermediate to the methylcitrate cycle. Moreover, it seems that the presence of a mixture of VFAs switches electron sinking from CO2 fixation to H2 and isoleucine production.

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“…Acetolactate synthase activity was examined as described previously ( Muhitch, 1988 ). The acetolactate produced after 1 h was assayed at a single end point by conversion to acetoin, which was detected by the reaction of Westerfeldt ( Westerfeldt, 1945 ) and quantified through the use of a standard curve after subtraction of the acetoin produced without substrate. Acetoin content was then normalized by the protein content.…”

Section: Methodsmentioning

confidence: 99%

Analysis of the Involvement of the Isoleucine Biosynthesis Pathway in Photoheterotrophic Metabolism of Rhodospirillum rubrum

et al. 2021

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Purple non-sulfur bacteria (PNSB) are recognized as a highly versatile group of bacteria that assimilate a broad range of carbon sources. Growing heterotrophically, PNSB such as Rhodospirillum rubrum (Rs. rubrum) generate reduced equivalents that are used for biomass production. However, under photoheterotrophic conditions, more reduced electron carriers than required to produce biomass are generated. The excess of reduced equivalents still needs to be oxidized for the metabolism to optimally operate. These metabolic reactions are known as electron sinks. Most PNSB rely on the CO2-fixing Calvin cycle and H2 production to oxidize these reduced equivalents. In addition to these well-described electron sinks, the involvement of some pathways, such as polyhydroxyalkanoate (PHA) biosynthesis, in redox poise is still controversial and requires further studies. Among them, isoleucine biosynthesis has been recently highlighted as one of these potential pathways. Here, we explore the role of isoleucine biosynthesis in Rs. rubrum. Our results demonstrate that the isoleucine content is higher under illuminated conditions and that submitting Rs. rubrum to light stress further increases this phenomenon. Moreover, we explore the production of (p)ppGpp in Rs. rubrum and its potential link with light stress. We further demonstrate that a fully functional isoleucine biosynthesis pathway could be an important feature for the onset of Rs. rubrum growth under photoheterotrophic conditions even in the presence of an exogenous isoleucine source. Altogether, our data suggest that isoleucine biosynthesis could play a key role in redox homeostasis.

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A Colorimetric Determination of Blood Acetoin

Cited by 638 publications

References 16 publications

Effect of variation ofKlebsiella pneumoniae acetolactate synthase expression on metabolic flux redistribution inEscherichia coli

Effect of variation ofKlebsiella pneumoniae acetolactate synthase expression on metabolic flux redistribution inEscherichia coli

Effects of Mixing Volatile Fatty Acids as Carbon Sources on Rhodospirillum rubrum Carbon Metabolism and Redox Balance Mechanisms

Analysis of the Involvement of the Isoleucine Biosynthesis Pathway in Photoheterotrophic Metabolism of Rhodospirillum rubrum

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