Thermodynamics and Structural Features of the Yeast<i>Saccharomyces cerevisiae</i>External Invertase Isoforms in Guanidinium-chloride Solutions

Andjelković, Uroš; Lah, Jurij

doi:10.1021/jf103441p

Cited by 10 publications

(4 citation statements)

References 30 publications

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“…Invertase 2 from A. terreus was more thermostable than invertases from A. ochraceus and Bacillus macerans , which exhibited half‐life of 1 hr when incubated at 60 and 55°, respectively (Guimarães et al, ; Park, Oh, & Yun, ). Natural thermostability can be due to a compact protein structure or due to high glycosylation as observed for invertase from Saccharomyces cerevisiae (Andjelković & Lah, ).…”

Section: Resultsmentioning

confidence: 99%

Purification and characterization of an invertase and a transfructosylase from Aspergillus terreus

et al. 2018

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Two intracellular invertases from Aspergillus terreus were purified and characterized. Invertase 1 has been shown maximal activity at pH 3.0 and optimum temperature at 60°C, while invertase 2 presented maximal activity at pH 4.6 and optimum temperature at 55°C. Km of invertase 1 and 2 for sucrose were 9.8 and 6.2 mM, respectively. Invertase 2 showed high thermostability presenting half‐life of 44, 14.9, and 2 hr at 40, 55, and 60°C, respectively. Gel filtration and SDS‐PAGE analysis revealed that invertase 2 has a dimeric structure composed by two monomers of 32 kDa. Fructose was a competitive inhibitor for invertase 2 presenting Ki of 61.5 mM. Besides hydrolytic activity, invertase 2 also showed transfructosylating activity producing fructooligosaccharides. The enzymes have potential to be applied in food industry since its product, inverted sugar, is used in candies and syrup production, while fructooligosacharides are prebiotics, low calorie and noncariogenic sweeteners. Practical applications Enzymes invertases hydrolyze sucrose in glucose and fructose producing inverted sugar. This product is widely used in food industry to produce jams, syrups, and fondant fillings for chocolates. Besides hydrolysis activity, some invertases can also present transfructosylating activity producing fructooligosaccharides. These oligosaccharides can act as prebiotic, providing beneficial health effects and can be used as low calorie and noncariogenic sweeteners. Aspergillus terreus produced two intracellular enzymes, one exclusively invertase, and the other, invertase and transfructosylase. The enzymes were purified and characterized in this study. This is an important step on screening new enzymes able to produce inverted sugar and fructooligosaccharides from sucrose.

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Section: Resultsmentioning

confidence: 99%

Purification and characterization of an invertase and a transfructosylase from Aspergillus terreus

et al. 2018

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“…This result is higher than described for enzymes from A. terreus (11.3%) (Giraldo et al., 2014), A. phoenicis (12.23%) (Rustiguel et al., 2011), A. niger (17%) (Nguyen et al., 2005), Pycnoporus sanguineus (24%) (Quiroga et al., 1995), A. ochraceus (30%) (Ghosh et al., 2001) and (41%) (Guimarães et al., 2007), T. pretoriensis (50%) (Oda & Tonomura, 1994), A. niveus (53.5%) (Guimarães et al., 2009), Schizosaccharomyces pombe (67%) (Moreno et al., 1990), and A. sojae (68.92%) (Lincoln & More, 2018). Andjelković and Lah (2011) described that the high level of glycosylation increases thermal stability, resistance to the action of proteases and enzymatic solubility of invertases produced by Saccharomyces cerevisiae . These characteristics are essential for enzyme application in industrial processes.…”

Section: Discussionmentioning

confidence: 99%

Cunninghamella echinulata PA3S12MM invertase: Biochemical characterization of a promiscuous enzyme

et al. 2021

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The Cunninghamella echinulata PA3S12MM fungus is a great producer of invertases in a growth medium supplemented by apple peels. The enzyme was purified 4.5 times after two chromatographic processes, and it presented a relative molecular mass of 89.2 kDa. The invertase reached maximum activity at pH of 6 and at 60°C, in addition to presenting stability in alkaline pH and thermal activation at 50°C. The enzymatic activity increased in the presence of Mn 2+ and dithiothreitol (DTT), while Cu 2+ and Z 2+ ions inhibited it. Also, DTT showed to protect enzymatic activity. The apparent values for K m , V máx , and K cat for the sucrose hydrolysis were, respectively, 173.8 mmol/L, 908.7 mmol/L min −1 , and 1,388.79 s −1 . The carbohydrate content was of 83.13%. The invertase presented hydrolytic activity over different types of glycosidic bonds, such as α1 ↔ 2β (sucrose), α1 → 4 (polygalacturonic acid), α1 → 4 and α1 → 2 (pectin), and α1 ↔ 1 (trehalose), indicating that the enzyme is multifunctional.Thus, the biochemical properties showed by the C. echinulata PA3S12MM suggest a broad industrial application, such as in the biomass hydrolysis or in the food industry.

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“…These forms exhibit the same catalytic properties (e.g., Km for sucrose) and the pH optimum [14]. At the other hand, their pI values, thermal stability, and chemical reactivity toward epoxy‐activated Eupergit C beads for enzyme immobilization are different [14], as well as structural features and thermodynamic stability in guanidinium‐chloride solutions [31]. These differences correlate with surface negative charge density and the content of phosphate groups bound to high‐mannose N ‐glycans [28].…”

Section: Introductionmentioning

confidence: 99%

Increased yield of enzymatic synthesis by chromatographic selection of different N‐glycoforms of yeast invertase

et al. 2020

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Invertases are glycosidases applied for synthesis of alkyl glycosides that are important and effective surfactants. Stability of invertases in the environment with increased content of organic solvent is crucial for increase of productivity of glycosidases. Their stability is significantly influenced by N-glycosylation. However, yeast N-glycosylation pathways may synthesize plethora of N-glycan structures. A total natural crude mixture of invertase glycoforms (EINV) extracted from Saccharomyces cerevisiae was subfractionated by anionexchange chromatography on industrial monolithic supports to obtain different glycoforms (EINV1-EINV3). Separated glycoforms exhibited different stabilities in water-alcohol solutions that are in direct correlation with the amount of phosphate bound to N-glycans.Observed differences in stability of different invertase glycoforms were used to improve productivity of methyl β-D-fructofuranoside (MF) synthesis. The efficiency and yield of MF synthesis was improved more than 50% when the most stabile glycoform bearing the lowest amount of phosphorylated N-glycans is selected and utilized. These data underline the importance of analysis of glycan structures attached to glycoproteins, demonstrates different impact of N-glycans on the surface charge and enzyme stability in regard to particular reaction environment, and provide a platform for improvement of yield of industrial enzymatic synthesis by chromatographic selection of glycoforms on monolithic supports.

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Thermodynamics and Structural Features of the YeastSaccharomyces cerevisiaeExternal Invertase Isoforms in Guanidinium-chloride Solutions

Cited by 10 publications

References 30 publications

Purification and characterization of an invertase and a transfructosylase from Aspergillus terreus

Purification and characterization of an invertase and a transfructosylase from Aspergillus terreus

Cunninghamella echinulata PA3S12MM invertase: Biochemical characterization of a promiscuous enzyme

Increased yield of enzymatic synthesis by chromatographic selection of different N‐glycoforms of yeast invertase

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