Stabilization of α‐amylase by chemical modification with carboxymethylcellulose

Villalonga, Reynaldo; Gómez, Leissy; Ramírez, Héctor L.; Villalonga, María L.

doi:10.1002/(sici)1097-4660(199907)74:7<635::aid-jctb85>3.3.co;2-d

Cited by 9 publications

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“…The attached polysaccharide molecules could also avoid the occurrence of protein aggregation processes in the enzyme at high temperatures, which play an important role in the thermal inactivation mechanism of trypsin (Moskvichyov et al 1986). It should be noted that the conjugates could be also thermostabilized by the formation of new intramolecular salt bridges, similarly to other enzymes chemically modified with ionic polymers (Villalonga et al 1999Go´mez et al 2001). However, the lower thermal stability showed by the CMC-trypsin adduct suggests a lower degree of both covalent and non-covalent intramolecular interactions between the globular protein structure and the polymeric chains.…”

Section: Discussionmentioning

confidence: 95%

“…This selection has been inspired by naturally occurring glycoenzymes, in which the sugar residues play an important role in the stability properties of these proteins (Wang et al 1996). In this regard, the covalent attachment of ionic and non-ionic polysaccharides such as chitosan Darias & Villalonga 2001), pectin , dextran (Blomhoff & Christensen 1983;Srivastava 1991), polymerized sucrose , carboxymethylcellulose (Villalonga et al 1999;Ramı´rez et al 2002), mannan (Masa´rova´et al 2001), alginate (Go´mez et al 2001) and cyclodextrin-grafted polysaccharides (Darias et al 2002;Villalonga et al 2003a) has been reported as a useful tool for increasing stability of enzymes.…”

Section: Introductionmentioning

confidence: 99%

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Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides

Villalonga

Mariniello

et al. 2006

World J Microbiol Biotechnol

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Dextran (MW=7.2x10(4)), carboxymethylcellulose (MW=2.5x10(4), substitution degree=0.7) and Ficoll (MW=6.9x10(4)) were derivatized with 1,4-diaminobutane and covalently attached to bovine pancreatic trypsin through a transglutaminase-catalysed reaction. The conjugates contained an average of 0.7-1.8 mol of polymers per mol of protein, and retained about 61-82% of the original esterolytic activity of trypsin. The optimum pH for trypsin was shifted to alkaline values after enzymatic glycosidation. The thermostability of the polymer-enzyme complexes was increased in about 13.7-50.0 degrees C over 10 min incubation. The prepared conjugates were also more stable against thermal incubation at different temperatures ranging from 50 degrees C to 60 degrees C. In comparison with native trypsin, the enzyme-polymer complexes were about 22- to 48-fold more resistant to autolytic degradation at pH 9.0. Transglutaminase-catalysed glycosidation also protected trypsin against denaturation in surfactant media, with 9- to 68-fold increased half-life times in the presence of 0.3% (w/v) sodium dodecylsulfate

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Section: Discussionmentioning

confidence: 95%

Section: Introductionmentioning

confidence: 99%

Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides

Villalonga

Mariniello

et al. 2006

World J Microbiol Biotechnol

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“…Recently, we reported that the covalent attachment of carboxymethylcellulose to porcine pancreatic α-amylase (PPA, EC 3.2.1.1) and trypsin improved the stability for these enzymes under various denaturing conditions [7,8]. Similar stabilization was described for invertase by modification of its sugar chains with chitosan, a cationic polysaccharide [9].…”

Section: Introductionmentioning

confidence: 91%

“…Amylolytic activity for native and modified PPA was determined as described by VILLALONGA et al [7]. The total carbohydrates were determined by the phenol-sulphuric acid method [13] using glucose as a standard.…”

Section: Assaysmentioning

confidence: 99%

Modification ofα-Amylase by Sodium Alginate

2001

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Sodium alginate, activated by periodate oxidation, was covalently linked to porcine pancreatic α-amylase via reductive alkylation with NaBH 4 . The enzyme-polymer conjugate, purified by gel filtration on Fractogel EMD BioSEC (S), retained about 50% of the native specific amylolytic activity. The sugar content was estimated to be 712 mol of monosaccharides per mol of enzyme protein. An average of 11 amino groups out of 21 groups from α-amylase were modified with the polysaccharide. The functional stability was improved for α-amylase after conjugation with sodium alginate. In comparison with the native enzyme, the thermostability of α-amylase was increased by this modification. In addition, the stability in the range of pH 5.0-11.0 was improved for the modified enzyme. The conjugate was also more resistant to denaturation by 0.3% sodium dodecylsulphate, retaining about 10% of its initial activity after 120 min of incubation. The formation of stabilizing salt bridges in the protein surface of the α-amylase-polysaccharide complex was confirmed by FT-IR spectrometry. Attending to the results obtained, we conclude that the covalent attachment of the anionic polysaccharide sodium alginate to the enzymes might be a useful and non-expensive method for improving the stabilization of these biocatalysts under various denaturing conditions.

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“…In previous works, we reported the covalent conjugation of several enzymes with the anionic polysaccharide carboxymethylcellulose (CMC) [24][25][26], demonstrating the effectiveness of this strategy for improving the functional stability of these biocatalysts. The present paper deals with the chemical glycosidation of SOD with CMC through two different synthetic procedures and the effect of these transformations on the pharmacological and stability properties of the modified enzymes.…”

Section: Introductionmentioning

confidence: 99%

Improved Pharmacological Properties for Superoxide Dismutase Modified with Carboxymethycellulose

Domínguez¹,

Valdivia²,

Caballero³

et al. 2005

Journal of Bioactive and Compatible Polymers

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Superoxide dismutase (SOD) was chemically modified with carboxymethylcellulose through two different synthetic procedures: Reductive alkylation with the periodate-oxidized polymer (SOD-CMCox), and the formation of amide linkages through a carbodiimide catalyzed reaction (SODCMCedac). The SOD-CMCox and SOD-CMCedac conjugates contained about 1.8-1.2 mol of polymer per mol of protein, and retained 68-78% of the initial catalytic activity, respectively. The glycosidated enzymes were more resistant to inactivation with H 2 O 2 and their plasma half-life times were prolonged to 34.7 h -6.6 h when compared with 4.8 min for native SOD. The anti-inflammatory activity of the enzyme was 2-2.4 times increased after conjugation with the polymer.

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Stabilization of α‐amylase by chemical modification with carboxymethylcellulose

Cited by 9 publications

References 0 publications

Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides

Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides

Modification ofα-Amylase by Sodium Alginate

Improved Pharmacological Properties for Superoxide Dismutase Modified with Carboxymethycellulose

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