Limits...
Benzoic acid fermentation from starch and cellulose via a plant-like β-oxidation pathway in Streptomyces maritimus.

Noda S, Kitazono E, Tanaka T, Ogino C, Kondo A - Microb. Cell Fact. (2012)

Bottom Line: S. maritimus expressed β-glucosidase and high amylase-retaining activity compared to those of S. lividans and S. coelicolor.In addition, for effective benzoate production from cellulosic materials, we constructed endo-glucanase-secreting S. maritimus.To achieve further benzoate productivity, the L-phenylalanine availability needs to be improved in future.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe, 657-8501, Japan.

ABSTRACT

Background: Benzoic acid is one of the most useful aromatic compounds. Despite its versatility and simple structure, benzoic acid production using microbes has not been reported previously. Streptomyces are aerobic, Gram-positive, mycelia-forming soil bacteria, and are known to produce various kinds of antibiotics composed of many aromatic residues. S. maritimus possess a complex amino acid modification pathway and can serve as a new platform microbe to produce aromatic building-block compounds. In this study, we carried out benzoate fermentation using S. maritimus. In order to enhance benzoate productivity using cellulose as the carbon source, we constructed endo-glucanase secreting S. maritimus.

Results: After 4 days of cultivation using glucose, cellobiose, or starch as a carbon source, the maximal level of benzoate reached 257, 337, and 460 mg/l, respectively. S. maritimus expressed β-glucosidase and high amylase-retaining activity compared to those of S. lividans and S. coelicolor. In addition, for effective benzoate production from cellulosic materials, we constructed endo-glucanase-secreting S. maritimus. This transformant efficiently degraded the phosphoric acid swollen cellulose (PASC) and then produced 125 mg/l benzoate.

Conclusions: Wild-type S. maritimus produce benzoate via a plant-like β-oxidation pathway and can assimilate various carbon sources for benzoate production. In order to encourage cellulose degradation and improve benzoate productivity from cellulose, we constructed endo-glucanase-secreting S. maritimus. Using this transformant, we also demonstrated the direct fermentation of benzoate from cellulose. To achieve further benzoate productivity, the L-phenylalanine availability needs to be improved in future.

Show MeSH
(A) HPLC traces of benzoic acid analysis. Lane 1: Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution. Lane 2: The culture supernatant of S. maritimus. (B) UV spectra of benzoic acid. 2.5 mg/l Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution (dotted line), 4 mg/l benzoic acid fraction separated from the culture supernatant of S. maritimus by HPLC (solid line).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3403852&req=5

Figure 2: (A) HPLC traces of benzoic acid analysis. Lane 1: Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution. Lane 2: The culture supernatant of S. maritimus. (B) UV spectra of benzoic acid. 2.5 mg/l Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution (dotted line), 4 mg/l benzoic acid fraction separated from the culture supernatant of S. maritimus by HPLC (solid line).

Mentions: After cultivation of S. maritimus/WT using TSB medium, benzoate produced in S. maritimus was identified using a co-chromatography method and UV spectrophotometer [19,20]. Figure 2(A) shows a chromatogram of a standard sample of benzoic acid solution (Lane 1) and the culture supernatant of S. maritimus/WT containing produced benzoic acid (Lane 2). The peak of benzoic acid in the standard sample was found at about 8 min (Lane 1), which was also observed in the culture supernatant of S. maritimus/WT (Lane 2). The addition of L-phenylalanine into the initial culture medium increased the peak areas of benzoic acid (data not shown). The UV spectra of the benzoic acid fraction separated from the culture supernatant of S. maritimus/WT exhibit two major absorption peaks in the region of 190–300 nm, similar to the standard sample of benzoic acid (Figure 2(B)). In addition, we carried out MS spectra analysis. MS and tandem MS analysis show that the peak of benzoic acid dehydrogenated and decarboxylated was observed around m/z 121.10 and 77.00, respectively. These results demonstrated that S. maritimus/WT produces benzoate in the culture supernatant.


Benzoic acid fermentation from starch and cellulose via a plant-like β-oxidation pathway in Streptomyces maritimus.

Noda S, Kitazono E, Tanaka T, Ogino C, Kondo A - Microb. Cell Fact. (2012)

(A) HPLC traces of benzoic acid analysis. Lane 1: Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution. Lane 2: The culture supernatant of S. maritimus. (B) UV spectra of benzoic acid. 2.5 mg/l Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution (dotted line), 4 mg/l benzoic acid fraction separated from the culture supernatant of S. maritimus by HPLC (solid line).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3403852&req=5

Figure 2: (A) HPLC traces of benzoic acid analysis. Lane 1: Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution. Lane 2: The culture supernatant of S. maritimus. (B) UV spectra of benzoic acid. 2.5 mg/l Standard sample of benzoic acid in acetonitrile: phosphate buffer (50 mM, pH 2.5) (30:70) solution (dotted line), 4 mg/l benzoic acid fraction separated from the culture supernatant of S. maritimus by HPLC (solid line).
Mentions: After cultivation of S. maritimus/WT using TSB medium, benzoate produced in S. maritimus was identified using a co-chromatography method and UV spectrophotometer [19,20]. Figure 2(A) shows a chromatogram of a standard sample of benzoic acid solution (Lane 1) and the culture supernatant of S. maritimus/WT containing produced benzoic acid (Lane 2). The peak of benzoic acid in the standard sample was found at about 8 min (Lane 1), which was also observed in the culture supernatant of S. maritimus/WT (Lane 2). The addition of L-phenylalanine into the initial culture medium increased the peak areas of benzoic acid (data not shown). The UV spectra of the benzoic acid fraction separated from the culture supernatant of S. maritimus/WT exhibit two major absorption peaks in the region of 190–300 nm, similar to the standard sample of benzoic acid (Figure 2(B)). In addition, we carried out MS spectra analysis. MS and tandem MS analysis show that the peak of benzoic acid dehydrogenated and decarboxylated was observed around m/z 121.10 and 77.00, respectively. These results demonstrated that S. maritimus/WT produces benzoate in the culture supernatant.

Bottom Line: S. maritimus expressed β-glucosidase and high amylase-retaining activity compared to those of S. lividans and S. coelicolor.In addition, for effective benzoate production from cellulosic materials, we constructed endo-glucanase-secreting S. maritimus.To achieve further benzoate productivity, the L-phenylalanine availability needs to be improved in future.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe, 657-8501, Japan.

ABSTRACT

Background: Benzoic acid is one of the most useful aromatic compounds. Despite its versatility and simple structure, benzoic acid production using microbes has not been reported previously. Streptomyces are aerobic, Gram-positive, mycelia-forming soil bacteria, and are known to produce various kinds of antibiotics composed of many aromatic residues. S. maritimus possess a complex amino acid modification pathway and can serve as a new platform microbe to produce aromatic building-block compounds. In this study, we carried out benzoate fermentation using S. maritimus. In order to enhance benzoate productivity using cellulose as the carbon source, we constructed endo-glucanase secreting S. maritimus.

Results: After 4 days of cultivation using glucose, cellobiose, or starch as a carbon source, the maximal level of benzoate reached 257, 337, and 460 mg/l, respectively. S. maritimus expressed β-glucosidase and high amylase-retaining activity compared to those of S. lividans and S. coelicolor. In addition, for effective benzoate production from cellulosic materials, we constructed endo-glucanase-secreting S. maritimus. This transformant efficiently degraded the phosphoric acid swollen cellulose (PASC) and then produced 125 mg/l benzoate.

Conclusions: Wild-type S. maritimus produce benzoate via a plant-like β-oxidation pathway and can assimilate various carbon sources for benzoate production. In order to encourage cellulose degradation and improve benzoate productivity from cellulose, we constructed endo-glucanase-secreting S. maritimus. Using this transformant, we also demonstrated the direct fermentation of benzoate from cellulose. To achieve further benzoate productivity, the L-phenylalanine availability needs to be improved in future.

Show MeSH