Application of different feeding strategies in fed batch culture for pullulanase production using sago starch

Shankar, Ravi; Madihah,; Shaza, E M; Aswati, K O Nur; Suraini, A.A.; Kamarulzaman, K

doi:10.1016/j.carbpol.2013.10.031

Cited by 14 publications

(3 citation statements)

References 33 publications

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“…Pullulanase production has been well-studied in recent years because of its extensive application in the food and chemical fuel industries [ 24 ]. Pullulanase has been expressed in E. coli [ 23 ], B. subtilis [ 25 ], B. flavothermus [ 26 ], B. licheniformis [ 27 ], Brevibacillus choshinensis [ 24 ], Pichia pastoris [ 28 ] and Saccharomyces cerevisiae [ 29 ]. Expression levels are high in E. coli, but the use of E. coli is restricted in industrial food applications because of its potential pathogenicity.…”

Section: Introductionmentioning

confidence: 99%

High-level extracellular protein production in Bacillus subtilis using an optimized dual-promoter expression system

et al. 2017

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BackgroundWe recently constructed a Bacillus subtilis strain (CCTCC M 2016536) from which we had deleted the srfC, spoIIAC, nprE, aprE and amyE genes. This strain is capable of robust recombinant protein production and amenable to high-cell-density fermentation. Because the promoter is among the factors that influence the production of target proteins, optimization of the initial promoter, PamyQ from Bacillus amyloliquefaciens, should improve protein expression using this strain. This study was undertaken to develop a new, high-level expression system in B. subtilis CCTCC M 2016536.ResultsUsing the enzyme β-cyclodextrin glycosyltransferase (β-CGTase) as a reporter protein and B. subtilis CCTCC M 2016536 as the host, nine plasmids equipped with single promoters were screened using shake-flask cultivation. The plasmid containing the PamyQ′ promoter produced the greatest extracellular β-CGTase activity; 24.1 U/mL. Subsequently, six plasmids equipped with dual promoters were constructed and evaluated using this same method. The plasmid containing the dual promoter PHpaII–PamyQ′ produced the highest extracellular β-CGTase activity (30.5 U/mL) and was relatively glucose repressed. The dual promoter PHpaII–PamyQ′ also mediated substantial extracellular pullulanase (90.7 U/mL) and α-CGTase expression (9.5 U/mL) during shake-flask cultivation, demonstrating the general applicability of this system. Finally, the production of β-CGTase using the dual-promoter PHpaII–PamyQ′ system was investigated in a 3-L fermenter. Extracellular expression of β-CGTase reached 571.2 U/mL (2.5 mg/mL), demonstrating the potential of this system for use in industrial applications.ConclusionsThe dual-promoter PHpaII–PamyQ′ system was found to support superior expression of extracellular proteins in B. subtilis CCTCC M 2016536. This system appears generally applicable and is amenable to scale-up.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-017-0649-1) contains supplementary material, which is available to authorized users.

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Section: Introductionmentioning

confidence: 99%

High-level extracellular protein production in Bacillus subtilis using an optimized dual-promoter expression system

et al. 2017

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“…P ullulanase (EC 3.2.1.41), which catalyzes the hydrolysis of ␣-1,6-glucosidic linkages in pullulan, amylopectin, and the dextrins of amylopectin, is a well-known starch-debranching enzyme (1). It belongs to the ␣-amylase family, which is identified as glycoside hydrolase family 13 in the CAZy database.…”

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confidence: 99%

Enhancing the Secretion Efficiency and Thermostability of a Bacillusderamificans Pullulanase Mutant (D437H/D503Y) by N-Terminal Domain Truncation

Duan

2015

Appl Environ Microbiol

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Pullulanase (EC 3.2.1.41), an important enzyme in the production of starch syrup, catalyzes the hydrolysis of ␣-1,6 glycosidic bonds in complex carbohydrates. A double mutant (DM; D437H/D503Y) form of Bacillus deramificans pullulanase was recently constructed to enhance the thermostability and catalytic efficiency of the enzyme (X. Duan, J. Chen, and J. Wu, Appl Environ Microbiol 79:4072-4077, 2013, http://dx.doi.org/10.1128/AEM.00457-13). In the present study, three N-terminally truncated variants of this DM that lack the CBM41 domain (DM-T1), the CBM41 and X25 domains (DM-T2), or the CBM41, X25, and X45 domains (DM-T3) were constructed. Upon expression, DM-T3 existed as inclusion bodies, while 72.8 and 74.8% of the total pullulanase activities of DM-T1 and DM-T2, respectively, were secreted into the medium. These activities are 2.8-and 2.9-fold that of the DM enzyme, respectively. The specific activities of DM-T1 and DM-T2 were 380.0 ؋ 10 8 and 449.3 ؋ 10 8 U · mol ؊1, respectively, which are 0.94-and 1.11-fold that of the DM enzyme. DM-T1 and DM-T2 retained 50% of their activity after incubation at 60°C for 203 and 160 h, respectively, which are 1.7-and 1.3-fold that of the DM enzyme. Kinetic studies showed that the K m values of DM-T1 and DM-T2 were 1.5-and 2.7-fold higher and the K cat /K m values were 11 and 50% lower, respectively, than those of the DM enzyme. Furthermore, DM-T1 and DM-T2 produced D-glucose contents of 95.0 and 94.1%, respectively, in a starch saccharification reaction, which are essentially identical to that produced by the DM enzyme (95%). The enhanced secretion and improved thermostability of the truncation mutant enzymes make them more suitable than the DM enzyme for industrial processes. P ullulanase (EC 3.2.1.41), which catalyzes the hydrolysis of ␣-1,6-glucosidic linkages in pullulan, amylopectin, and the dextrins of amylopectin, is a well-known starch-debranching enzyme (1). It belongs to the ␣-amylase family, which is identified as glycoside hydrolase family 13 in the CAZy database. When combined with ␣-amylase, ␤-amylase, glucoamylase, or cyclodextrin glucosyltransferase, pullulanase can be used in the production of glucose, fructose, maltose, cyclodextrins, and amylose (2, 3). The addition of pullulanase allows the reaction time to be reduced, the substrate concentration to be increased, and the conversion rate to be improved (4, 5). Recently, pullulanase has been used as a biocatalyst for the production of fuel ethanol, low-calorie beers, resistant starch, maltotriose, and other important products (6, 7). With this plethora of potential industrial applications, pullulanase production has attracted a great deal of attention in recent years (8,9).A large number of microorganisms, including mesophiles, thermophiles, and hyperthermophiles, have been shown to produce pullulanase (10, 11). Although numerous pullulanases have been identified and heterologously expressed (12)(13)(14), reports have shown that the secretion ratio of recombinant pullulanase is low. Low extracellular prod...

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“…Bence (2011) with comparison of two CFRs of 0.125 and 0.40 l/h showed that increase of dilution rate by the adding fed-batch bioreactor volume at a higher CFR (0.40 l/h) led to a significant improvement in glucose consumption and biosurfactant production by Bacillus subtilis. Shankar et al (2014) also found that the use of an ideal dilution rate can significantly supply high amounts of biomass and pullulanase synthesis by Bacillus flavothermus KWF-1 under constant feed rate mode.…”

Section: Analysis Of Influence Of Variables On the Cell Mass And Pigm...mentioning

confidence: 90%

Optimisation and kinetic studies on the production of intracellular canthaxanthin in fed-batch cultures of Dietzia natronolimnaea HS-1

Gharibzahedi

Razavi

Mousavi

2015

Quality Assurance and Safety of Crops & Foods

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A critical optimisation and kinetic study on the production improvement of carotenoid pigment extracted from Dietzia natronolimnaea HS-1 under fed-batch fermentation was conducted. High pressure liquid chromatography revealed that the most abundant pigment biosynthesised by this bacterium was canthaxanthin. The constant feeding rates (CFRs; 5.59-8.41 ml/h) for different concentrations of enzymatic hydrolysed molasses (EHM; 40.76-49.24 g/l) were optimised to achieve the highest cell biomass, total carotenoid and canthaxanthin amounts using a response surface method. Spectrophotometric and optimisation analysis showed that the maximum cell mass (17.79 g/l), total carotenoid (16.37 mg/l) and canthaxanthin (14.63 mg/l) can be obtained at 45.90 g/l EHM with a CFR of 6.40 ml/h. A very close agreement between experimental and predicted data was found. A substantial increase (123.4%) in production of canthaxanthin was achieved after 152 h fed-batch process under optimal CFR mode as compared with batch process. Moreover, the maximum specific growth rate and, the volumetric productivities and yields of cell mass, total carotenoid and canthaxanthin in fed-batch culture were significantly more than the batch one.

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Application of different feeding strategies in fed batch culture for pullulanase production using sago starch

Cited by 14 publications

References 33 publications

High-level extracellular protein production in Bacillus subtilis using an optimized dual-promoter expression system

High-level extracellular protein production in Bacillus subtilis using an optimized dual-promoter expression system

Enhancing the Secretion Efficiency and Thermostability of a Bacillusderamificans Pullulanase Mutant (D437H/D503Y) by N-Terminal Domain Truncation

Optimisation and kinetic studies on the production of intracellular canthaxanthin in fed-batch cultures of Dietzia natronolimnaea HS-1

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