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1.
Jinxin Zhang Karen M. J. Saerens Inge N. A. Van Bogaert Wim Soetaert 《Biotechnology letters》2011,33(12):2417-2423
The yeast Rhodotorula bogorensis produces sophorolipids of different structures to those produced by Candida bombicola. However, the yield is very low. To improve sophorolipid production by R. bogoriensis, vegetable oil was supplemented to the medium as a hydrophobic substrate: with rapeseed oil the sophorolipid yield was 1.26 g/l
but without oil was 0.33 g/l. Cultures with meadowfoam oil produced 0.77 g sophorolipids/l. Lipase-treated meadowfoam oil,
however, gave no significant increase in sophorolipid production. Possible explanations for the enhanced sophorolipid synthesis
are discussed. 相似文献
2.
【目的】槐糖脂是一类生物表面活性剂,不仅具有常规表面活性剂所具有的增溶、乳化、润湿、发泡、分散、降低表面张力等通用性能,且对环境的耐受性极强。熊蜂生假丝酵母(Starmerella bombicola)能够发酵生产槐糖脂,但槐糖脂具有酸型、内酯型和乙酰化型等不同类型,结构多样,难以分离。本文拟通过代谢工程改造,构建高产酸型槐糖脂的熊蜂生假丝酵母工程菌株。【方法】利用潮霉素抗性基因构建了标记基因重复利用系统Rec-six基因编辑系统,在此基础上将合成内酯型槐糖脂的关键基因——内酯酶基因SBLE敲除获得一株只产酸型槐糖脂的工程菌株Δsble,进一步同源过量表达葡萄糖基转移酶基因UGTB并敲除过氧化物酶体膜转运蛋白编码基因PXA1,构建了高产酸型槐糖脂的酵母工程菌。【结果】与出发菌株相比,重组熊蜂生假丝酵母发酵油酸能够合成单一的酸型槐糖脂,而不再合成内酯型槐糖脂,同时酸型槐糖脂的产量由20 g/L提高到44 g/L,提高了2.1倍。【结论】通过敲除PXA1、SBLE和过表达UGTB来改造熊蜂生假丝酵母,能够有效提高重组菌的酸型槐糖脂产量,为发酵法生产酸型槐糖脂奠定了基础。 相似文献
3.
4.
I. N. A. Van Bogaert S. Roelants D. Develter W. Soetaert 《Biotechnology letters》2010,32(10):1509-1514
Sophorolipids production by the yeast Candia bombicola is most favourable when glucose is used as a carbon source in combination with a hydrophobic carbon source such as a common
vegetable oil. Most vegetable oils are comprised of C16–C18 fatty acids, an ideal range for sophorolipid production. The use
of oils with either shorter or longer fatty acids, such has coconut oil or meadowfoam oil, respectively, was evaluated. Such
oils did not contribute to enhanced sophorolipid production when compared to cultures run on glucose as the sole carbon source.
Moreover, a toxic effect of medium-chain fatty acids towards stationary C. bombicola cells was demonstrated. 相似文献
5.
Synthesis of medium-chain sophorolipids by Candida bombicola is a challenging objective. One of the difficulties is that the obtained sophorolipids always represent a mixture of medium-chain
and native de novo formed or long-chain sophorolipids. The fatty acid moiety of de novo sophorolipids is derived from the
de novo synthesis of fatty acids. Fatty acid synthesis can be blocked by the antifungal agent cerulenin, an inhibitor if the
fatty acid synthase (FAS) complex acting on the β-ketoacyl thioester synthetase reaction. The toxic effect of cerulenin on
C. bombicola was evaluated and 20 mg/ml was added in the stationary growth phase. No de novo formed sophorolipids were observed when the
cells were cultured on merely glucose. Also when the hydrophilic substrate, 1,12-dodedanediol, was added, no de novo formed
sophorolipids were detected, leading to a reduced complexity of the sophorolipid mixture. 相似文献
6.
Inge N.A. Van Bogaert Julia Sabirova Dirk Develter Wim Soetaert & Erick J. Vandamme 《FEMS yeast research》2009,9(4):610-617
The nonpathogenic yeast Candida bombicola synthesizes sophorolipids. These biosurfactants are composed of the disaccharide sophorose linked to a long-chain hydroxy fatty acid and have potential applications in the food, pharmaceutical, cosmetic and cleaning industries. In order to expand the range of application, a shift of the fatty acid moiety towards medium-chain lengths would be recommendable. However, the synthesis of medium-chain sophorolipids by C. bombicola is a challenging objective. First of all, these sophorolipids can only be obtained by fermentations on unconventional carbon sources, which often have a toxic effect on the cells. Furthermore, medium-chain substrates are partially metabolized in the β-oxidation pathway. In order to redirect unconventional substrates towards sophorolipid synthesis, the β-oxidation pathway was blocked on the genome level by knocking out the multifunctional enzyme type 2 (MFE-2) gene. The total gene sequence of the C. bombicola MFE-2 (6033 bp) was cloned (GenBank accession number EU371724 ), and the obtained nucleotide sequence was used to construct a knock-out cassette. Several knock-out mutants with the correct geno- and phenotype were evaluated in a fermentation on 1-dodecanol. All mutants showed a 1.7–2.9 times higher production of sophorolipids, indicating that in those strains the substrate is redirected towards the sophorolipid synthesis. 相似文献
7.
Renjun Gao Mia Falkeborg Xuebing Xu Zheng Guo 《Applied microbiology and biotechnology》2013,97(3):1103-1111
To achieve high time–space efficiency for sophorolipid production with yeast Candida bombicola, a strategy of high cell density fermentation was employed. The approach consisted of two sequential stages: (1) the optimization of the carbon source and the nutrient concentration to achieve the maximal cell density and (2) the computer-aided adjustment of physical parameters and the controlled feeding of substrates for enhanced volumetric productivity. Both stages have been successfully implemented in a 10-L fermenter, where up to 80 g dry cell weight/L was obtained and a remarkably high volumetric productivity (> 200 g isolated sophorolipids/L/day) was achieved. Both the biomass and volumetric productivity were markedly higher than previously reported. Specifically, the high productivity of sophorolipids could be attained on a very short time scale (24 h), highlighting the industrial potential of the platform developed in this work. 相似文献
8.
Daniel HJ Otto RT Binder M Reuss M Syldatk C 《Applied microbiology and biotechnology》1999,51(1):40-45
In order to produce sophorolipids from whey, thereby lowering the lactose content and biological oxygen demand, a two-step
batch cultivation process was developed including medium sterilization by filtration. In the first step, whey was sterilized
by a combination of crossflow and sterile filtration. Because the sophorolipid-producing yeast Candida bombicola ATCC 22214 was not able to use lactose as a carbon source directly, the oleaginous yeast Cryptococcus curvatus ATCC 20509 was grown on deproteinized whey concentrates (DWC). With 1: 1 diluted DWC-20, lactose was consumed as the carbon
source and biomass (24 g/l dry weight content) as well as single-cell oil (SCO, 10 g/l) were produced. The cultivation broth
was disrupted with a glass bead mill and it served as medium for growth (29 g cell dry mass/l) and sophorolipid production
(12 g/l) of the yeast C. bombicola.
Received: 29 July 1998 / Received revision: 5 October 1998 / Accepted: 11 October 1998 相似文献
9.
Isabel A. Ribeiro M. Rosário Bronze Matilde F. Castro Maria H. L. Ribeiro 《Applied microbiology and biotechnology》2013,97(5):1875-1887
Microtiter plates were used as minireactors to study Starmerella bombicola growth and sophorolipid (SL) production. Compositional analysis of SL mixtures by liquid chromatography with electrospray ionization tandem mass spectrometry showed similar results on SLs produced using the laboratory scale (shake flask) and the microscale (24-well microtiter plates (MTP)) approach. MTP suitability on SL production was proven, being this approach, especially advantageous on SL screening. Several hydrophilic carbon sources, hydrophobic co-substrates and nitrogen sources were supplied to culture media, and their influence on SL production was evaluated. The selection of specific hydrophobic co-substrate and nitrogen sources influenced the ratio acidic/lactonic SLs. In fact, it was observed that the production of acidic C18:1 diacetylated hydroxy fatty acid SLs was favoured when culture media was supplied with avocado, argan, sweet almond and jojoba oil or when NaNO3 was supplied instead of urea. This last case was observed after 144 h of cultivation. A new SL, lactonic C18:3 hydroxy fatty acid diacetylated SL, was detected when borage and onagra oils were used individually as co-substrates. Overall results indicated the potential of the selective production of different and new sophorolipids by Starmerella bombicola based on the selection of carbon and nitrogen sources to culture media. 相似文献
10.
The effects of nitrogen sources on growth of sophorolipid-producing yeast, Wickerhamiella domercqiae var. sophorolipid CGMCC 1576 and on production and composition of sophorolipids were studied. Organic nitrogen sources are more favorable for
accumulation of biomass than inorganic ones. Presence of ammonium ion from different inorganic nitrogen sources (except NH4HCO3) greatly inhibited the production of lactonic sophorolipids. However, when organic nitrogen sources were used, lactonic sophorolipid
production was strongly increased. Production of crystalline lactonic sophorolipids from organic/inorganic nitrogen sources
was enhanced with the increase of pH value adjusted by sodium hydroxide or sodium citrate solution. Fourier-transform infrared
(FT-IR), gas chromatography mass spectrometry (GC-MS), high-performance liquid chromatography (HPLC), and mass spectra (MS)
were employed to compare the composition of sophorolipid mixture obtained from different nitrogen sources. More than 15 acidic
sophorolipid molecules and only 4 lactonic sophorolipid molecules were produced by using 1.27 g/l ammonium sulfate as nitrogen
source; they were separated by preparative HPLC and their structures were elucidated by MS. These results suggest extraordinary
regulatory effects of nitrogen source on growth and sophorolipid synthesis of W. domercqiae var. sophorolipid. 相似文献
11.
The yeast Candida bombicola (ATCC 22214) grown on primary carbon source glucose (100 g l−1) and secondary carbon, arachidonic acid (2 g l−1) produced mixture of sophorolipids up to 1.44 g l−1. The crude product was a heterogeneous mixture of sophorolipids, which are glycolipids of sophorose linked to the fatty acid
through glycosidic bond between ω and ω−1 carbon of arachidonic acid. The derived sophorolipids were isolated by silica gel
chromatography using dialysis tubing. The purified sophorolipids were characterized by ESI-MS and FT-IR. Acid hydrolysis of
the resolved sophorolipids were characterized by ESI-MS for the presence of 20-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid (20-HETE) and 19-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid (19-HETE), compounds of pronounced pharmaceutical importance. 相似文献
12.
Katarzyna Ciesielska Sophie L. K. W. Roelants Inge N. A. Van Bogaert Stijn De Waele Isabel Vandenberghe Sara Groeneboer Wim Soetaert Bart Devreese 《Applied microbiology and biotechnology》2016,100(22):9529-9541
We recently discovered a novel enzyme in the exoproteome of Starmerella bombicola, which is structurally related to Candida antarctica lipase A. A knockout strain for this enzyme does no longer produce lactonic sophorolipids, prompting us to believe that this protein is the missing S. bombicola lactone esterase (SBLE). SBLE catalyzes a rather unusual reaction, i.e., an intramolecular esterification (lactonization) of acidic sophorolipids in an aqueous environment, which raised questions about its activity and mode of action. Here, we report the heterologous production of this enzyme in Pichia pastoris and its purification in a two-step strategy. Purified recombinant SBLE (rSBLE) was used to perform HPLC and liquid chromatography mass spectrometry (LCMS)-based assays with different sophorolipid mixtures. We experimentally confirmed that SBLE is able to perform ring closure of acetylated acidic sophorolipids. This substrate was selected for rSBLE kinetic studies to estimate the apparent values of K m . We established that rSBLE displays optimal activity in the pH range of 3.5 to 6 and has an optimal temperature in the range of 20 to 50 °C. Additionally, we generated a rSBLE mutant through site-directed mutagenesis of Ser194 in the predicted active site pocket and show that this mutant is lacking the ability to lactonize sophorolipids. We therefore propose that SBLE operates via the common serine hydrolase mechanism in which the catalytic serine residue is assisted by a His/Asp pair. 相似文献
13.
Wadekar SD Kale SB Lali AM Bhowmick DN Pratap AP 《Preparative biochemistry & biotechnology》2012,42(2):125-142
Biosurfactants are microbially synthesized surfactants that are environmental friendly due to low toxicity. Sophorolipid is one of the simplest biosurfactants with well-defined structure produced by Starmerella bombicola(ATCC 22214) on glucose and vegetable oil as the carbon source. The raw material cost accounts for 10-30% of the overall cost. Glycerol is readily available from a commercial fat-splitting process as sweetwater at a very low cost. Sophorolipids was synthesized using glycerol and sweetwater as a cost-effective carbon source. The glycerol was further replaced with sweetwater as a source of glycerol. Optimum glycerol concentration was 15% w/v with 10% w/v sunflower oil, giving 6.6 g/L of sophorolipids. The crude sophorolipid contains two major components; both of them were lactonic sophorolipids as analyzed by reverse-phase high-performance liquid chromatography (RP-HPLC), liquid chromatography-mass spectroscopy (LC-MS), and nuclear magnetic resonance ((1)H-NMR). 相似文献
14.
《Experimental mycology》1990,14(4):405-415
The relationship between β-linked disaccharide (cellobiose, sophorose) utilization and cellulase, particularly cellobiohydrolase I (CBH I) synthesis by Trichoderma reesei, was investigated. During growth on cellobiose and sophorose as carbon sources in batch as well as resting-cell culture, only sophorose induced cellulase formation. In the latter experiments, sophorose was utilized at a much lower rate than cellobiose, and the more cellulase produced, the lower its rate of utilization. Cellobiose and sophorose were utilized by the fungus mainly via hydrolysis by the cell wall- and cell membrane-bound β-glucosidase. Addition of sophorose to T. reesei growing on cellulose did not further stimulate cellulase synthesis, and addition of cellobiose was inhibitory. Cellobiose, however, promoted cellulase formation in both batch and resting cell cultures, when its hydrolysis by β-glucosidase was inhibited by nojirimycin. No cellulase formation was observed when the uptake of glucose (produced from cellobiose by β-glucosidase) was inhibited by 3-O-methylglucoside. Cellodextrins (C2 to C6) promoted formation of low levels of cellobiohydrolase I in indirect proportion to their rate of hydrolysis by β-glucosidase. Studies on the uptake of [3H]cellobiose, [3H]sophorose, and [14C]glucose in the presence of inhibitors of β-glucosidase (nojirimycin) and glucose transport (3-O-methylglucoside) show that glucose transport occurs at a much higher rate than disaccharide hydrolysis. Extracellular disaccharide hydrolysis accounts for at least 95% of their metabolism. The presence of an uptake system for cellobiose was established by demonstrating the presence of intracellular labeled [3H]cellobiose in T. reesei after its extracellular supply. The data are consistent with induction of cellulase and particularly CBH I formation in T. reesei by β-linked disaccharides under conditions where their uptake is favored at the expense of extracellular hydrolysis. 相似文献
15.
A key barrier to market penetration for sophorolipid biosurfactants is the ability to improve productivity and utilize alternative feedstocks to reduce the cost of production. To do this, a suitable screening tool is required that is able to model the interactions between media components and alter conditions to maximize productivity. In the following work, a central composite design is applied to analyse the effects of altering glucose, rapeseed oil, corn steep liquor and ammonium sulphate concentrations on sophorolipid production with Starmerella bombicola ATCC 222144 after 168 h. Sophorolipid production was analysed using standard least squares regression and the findings related to the growth (OD600) and broth conditions (glucose, glycerol and oil concentration). An optimum media composition was found that was capable of producing 39.5 g l–1 sophorolipid. Nitrogen and rapeseed oil sources were found to be significant, linked to their role in growth and substrate supply respectively. Glucose did not demonstrate a significant effect on production despite its importance to biosynthesis and its depletion in the broth within 96 h, instead being replaced by glycerol (via triglyceride breakdown) as the hydrophilic carbon source at the point of glucose depletion. A large dataset was obtained, and a regression model with applications towards substrate screening and process optimisation developed. 相似文献
16.
17.
A.-M. Davila R. Marchal J.-P. Vandecasteele 《Applied microbiology and biotechnology》1997,47(5):496-501
Sophorose lipid production by Candida bombicola is a two-step process where sophorose lipids are mainly produced after a first stage of growth, ending because of nitrogen
limitation. The influence of the following parameters was individually studied for both the stages of growth and of product
formation with respect to final sophorose lipid production performance: pH, temperature and carbon source. Glucose and rapeseed
ethyl esters were supplied individually or as a dual carbon source. The lipidic substrate was added by continuous feeding.
It was found that supplying both carbon sources during the production step was crucial for obtaining a high production performance
ranging from 250 g l−1 to 300 g l−1 or more. Controlling the feeding of rapeseed ethyl esters to avoid inhibition by fatty acids was essential for a successful
scale-up of the fermentation on the industrial scale. The conditions of substrate feeding markedly affected the composition
of the mixture of sophorose lipids produced, namely the extent of acetylation of the sophorose moieties and distribution of
the acidic and lactonic forms. The results suggest that the physiological role of sophorose lipid production is related to
the regulation of energy metabolism.
Received: 26 June 1996 / Received revision: 12 December 1996 / Accepted: 15 December 1996 相似文献
18.
Qingxin Zhou Jintao Xu Yanbo Kou Xinxing Lv Xi Zhang Guolei Zhao Weixin Zhang Guanjun Chen Weifeng Liu 《Eukaryotic cell》2012,11(11):1371-1381
Appropriate perception of cellulose outside the cell by transforming it into an intracellular signal ensures the rapid production of cellulases by cellulolytic Hypocrea jecorina. The major extracellular β-glucosidase BglI (CEL3a) has been shown to contribute to the efficient induction of cellulase genes. Multiple β-glucosidases belonging to glycosyl hydrolase (GH) family 3 and 1, however, exist in H. jecorina. Here we demonstrated that CEL1b, like CEL1a, was an intracellular β-glucosidase displaying in vitro transglycosylation activity. We then found evidence that these two major intracellular β-glucosidases were involved in the rapid induction of cellulase genes by insoluble cellulose. Deletion of cel1a and cel1b significantly compromised the efficient gene expression of the major cellulase gene, cbh1. Simultaneous absence of BglI, CEL1a, and CEL1b caused the induction of the cellulase gene by cellulose to further deteriorate. The induction defect, however, was not observed with cellobiose. The absence of the three β-glucosidases, rather, facilitated the induced synthesis of cellulase on cellobiose. Furthermore, addition of cellobiose restored the productive induction on cellulose in the deletion strains. The results indicate that the three β-glucosidases may not participate in transforming cellobiose beyond hydrolysis to provoke cellulase formation in H. jecorina. They may otherwise contribute to the accumulation of cellobiose from cellulose as inducing signals. 相似文献
19.
Mehdi Dashtban Robert Buchkowski Wensheng Qin 《International Journal of Biochemistry and Molecular Biology》2011,2(3):274-286
The ascomycete Hypocrea jecorina, an industrial (hemi)cellulase producer, can efficiently degrade plant polysaccharides. At present, the biology underlying cellulase hyperproduction of T. reesei, and the conditions for the enzyme induction, are not completely understood. In the current study, three different strains of T. reesei, including QM6a (wild-type), and mutants QM9414 and RUT-C30, were grown on 7 soluble and 7 insoluble carbon sources, with the later group including 4 pure polysaccharides and 3 lignocelluloses. Time course experiments showed that maximum cellulase activity of QM6a and QM9414 strains, for the majority of tested carbon sources, occurred at 120 hrs, while RUT-C30 had the greatest cellulase activity around 72 hrs. Maximum cellulase production was observed to be 0.035, 0.42 and 0.33 µmol glucose equivalents using microcrystalline celluloses for QM6a, QM9414, and RUTC-30, respectively. Increased cellulase production was positively correlated in QM9414 and negatively correlated in RUT-C30 with ability to grow on microcrystalline cellulose. 相似文献
20.
The ability of Candida bombicola ATCC 22214 to produce sophorolipids using Turkish corn oil and honey was investigated. Shake flask experiments were carried out both with and without the addition of glucose as the second carbon source. The organism could produce sophorolipids under both conditions but higher production was obtained when corn oil was combined with glucose. The 3 L bioreactor was first operated in batch mode, using both corn oil and glucose. When all the glucose was consumed, 1/4th of the broth was pumped out and was replaced by freshly prepared medium containing 10 % [w/v] of cheap market honey as the sole carbon source. Feed was comprised of corn oil. High concentrations of sophorolipids (> 400 g/L) were produced. The crude products obtained from the batch cultivation could be solidified as very light brown solids when unused oil was removed by hexane, while the products of the two‐stage cultivation remained as viscous, honey‐like liquids after identical treatments. 相似文献