Raw starch degradation by the non-raw starch-adsorbing bacterial alpha amylase of Bacillus sp. IMD 434

Hamilton, Lynn; Kelly, Catherine T.; Fogarty, William M.

doi:10.1016/s0008-6215(98)00300-0

Cited by 44 publications

(24 citation statements)

References 9 publications

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“…The catalytic domain consisting of the A and B domains is normally located at the N-terminus. These domains can be recovered as separate fragments by proteolytic digestion using either trypsin or other proteases (Desseaux et al 1991;Kim & Kho 1993;Ferey-Roux et al 1998;Hamilton et al 1998;Khajeh et al 2006). Here we have shown the two fragments obtained during tryptic digestion of Sfamy, which is in agreement with the previous studies.…”

Section: Discussionmentioning

confidence: 99%

Proteolysis of α-amylase from Saccharomycopsis fibuligera: characterization of digestion products

et al. 2008

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α-Amylase from Saccharomycopsis fibuligera R-64 was successfully purified by butyl Toyopearl hydrophobic interaction chromatography, followed by Sephadex G-25 size exclusion and DEAE Toyopearl anion exchange chromatography. The enzyme has a molecular mass of 54 kDa, as judged by SDS PAGE analysis. Upon tryptic digestion, two major fragments with relative molecular masses of 39 kDa and 10 kDa, which resemble the A/B and C-terminal domains in the homologous Taka-amylase, were obtained and were successfully separated with the Sephadex G-50 size exclusion column. The 39-kDa fragment demonstrated a similar amylolytic activity to that of the undigested enzyme. However, it was found that the Km value of the 39-kDa fragment was about two-times higher than that of the undigested enzyme. Moreover, thermostability studies showed a lower half-life time for the 39-kDa fragment. These findings suggest that the 39-kDa fragment is the catalytic domain, while the 10-kDa fragment is the C-terminal one, which plays a role in thermostability and starch binding. Although the undigested enzyme is able to act on raw starches at room temperature, with maize starches as the best substrate, neither the undigested enzyme nor the fragments adsorb the tested raw starches. These results propose Saccharomycopsis fibuligera α-amylase as a raw starch-digesting but not adsorbing amylase, with a similar domain organization to that of Taka-amylase A.

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Section: Discussionmentioning

confidence: 99%

Proteolysis of α-amylase from Saccharomycopsis fibuligera: characterization of digestion products

et al. 2008

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“…β-Glucosidase exhibited Michaelis Menten type kinetics and K m and V max were found to be 3.11 mM and 20.83 U/mg respectively. K m is a measure of affinity of an enzyme for a substrate and low values of K m indicate high affinity of the enzyme for the substrate (Hamilton et al, 1998). Seidle et al (2004) calculated K m and V max of 1 mM and 40 U/mg respectively for pNPG hydrolysis with a β-glucosidase from A. niger.…”

Section: Enzyme Characterizationmentioning

confidence: 99%

Metabolic engineering and thermodynamic characterization of an extracellular -glucosidase produced by Aspergillus niger

Zahoor¹,

Javed²,

Aftab³

et al. 2011

Afr. J. Biotechnol.

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The production of an extracellular β-glucosidase by Aspergillus niger NRRL 599 was optimized using submerged fermentation technique. Effect of different media, different carbon sources, initial pH of the fermentation medium, temperature, incubation period and inoculum size on the production of β-glucosidase enzyme was investigated. A. niger NRRL 599 produced maximum extracellular β-glucosidase (4.48 U/mg) in Eggins and Pugh medium with 1% wheat bran (w/v) at pH 5.5 inoculated with 4% conidial suspension after 96 h of incubation at 30°C. Purified β-glucosidase gave K m and V max values of 3.11 mM and 20.83 U/mg respectively for pNPG hydrolysis. The enzyme was optimally active at pH 4.8 and at temperature of 60°C. Thermodynamic parameters, E a , ∆H and ∆S were found to be 52.17 KJ/mol, 49.90 J/mol.K and -71.69 KJ/mol, respectively. The pKa 1 and pKa 2 of ionizable groups of active site residues involved in V max were calculated to be 4.1 and 6.0 respectively.

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“…There was actually a three fold enhancement in the enzyme activity at this level of vitamin B12 supplementation. This clearly indicates that not only the quality of the nutrient but also its observations that high temperature inactivation may be due to incorrect conformation due to hydrolysis of the peptide chain, destruction of amino acid or aggregation, were made by earlier workers [5,24].…”

Section: Discussionmentioning

confidence: 58%

Production and partial purification of α-amylase from Pseudomonas sp. 2 under solid-state fermentation

Varalakshmi¹,

Pallavi²,

Banerjee³

et al. 2012

Turk J Bioch

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Purpose: The purpose of the present study was to isolate Pseudomonas sp. 2 from rhizosphere and screen for best amylase producer and to check for its suitability for industrial applications. Methods: Four strains of Pseudomonas were isolated from rhizosphere soil, by serial dilution method, of which Pseudomonas sp. 2 gave best results to α-amylase production. This strain was used to determine the enzyme production by SSF on wheat bran. Results and conclusion: The optimal enzyme production was obtained at 37 ºC and pH 7.5. Vitamins B 12 (50 mg) followed by Pyridoxine (50 mg) were found to be enhancing the enzyme production. Amino acids cysteine (30 mg), tyrosine (40 mg) and alanine (30 mg) were stimulating the enzyme production. The enzyme was partially purified by ammonium sulphate precipitation, and was further purified by ion exchange DEAE-cellulose column Chromatography. The purity of the enzyme was 45.2 fold greater than the crude enzyme. The molecular weight of the enzyme was found to be ~62 kDa by SDS-PAGE. This is the first report of any Pseudomonas sp. from rhizosphere being used for amylase production. The purified enzyme was not thermostable but it was acid tolerant making it suitable for industrial application. Key words: Pseudomonas sp. 2, α-amylase, SSF, Vitamins, Amino acids, Partial purification. Conflict of interest:The authors declare no conflicts of interest of any kind. ÖZETAmaç: Çalışamanın amacı Rizofer'den Pseudomonas sp. 2 bakterisini izole etmek, bundan en iyi amilaz üretimini bulmak ve bunun endüstriyel aplikasyonlara uygunluğunu kontrol etmektir. Metot: Seri dilüsyon metodu ile 4 tane Pseudomonas suju izole eidilmiştir. Bunların arasından Pseudomonas sp. 2 en iyi α-amilaz üretimini vermiştir. Bu suj, buğday kepeğinde SSF ile enzim üretiminde kullanılmıştır. Sonuçlar ve Tartışma: Optimum enzim üretimi 37 ºC and pH 7.5'da elde edilmiştir. Vitamin B 12 (50 mg) ve takiben piridoksin (50 mg) muamelelerinin enzim üretimini arttırdığı bulunmuştur. Sistein (30 mg), ıirozin (40 mg) ve alanin (30 mg) amino asitlerinin de enzim üretimini arttırdığı gözlenmiştir. Enzim, amonyum sülfat presipitasyonu ve DEAE-selüloz kolon kromatografisi ile kısmen saflaştırılmıştır. Saflaştırma verimi, ham enzimden 45.2 kez fazladır. SDS-PAGE kullanılarak enzimin molekül ağırlığı ~62 kDa bulunmuştur. Bu çalışma, Rizosfer'den elde edilen Pseudomonas sp.'dan ilk defa amilaz elde edilmesi ve kullanılmasını sunmaktadır. Kısmen saflaştırılmış enzim termostabil değildir, fakat aside dayanıklıdır ki bu enzimi endüstriyel uygulamalar için uygun hale getirmektedir. Anahtar Kelimeler: Pseudomonas sp. 2, α-amilaz, SSF, Vitaminler, Aminoasitler, Kısmi saflaştırma. Çıkar çatışması: Yazarlar hiçbir çıkar çatışması bulunmadığını beyan ederler.

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Raw starch degradation by the non-raw starch-adsorbing bacterial alpha amylase of Bacillus sp. IMD 434

Cited by 44 publications

References 9 publications

Proteolysis of α-amylase from Saccharomycopsis fibuligera: characterization of digestion products

Proteolysis of α-amylase from Saccharomycopsis fibuligera: characterization of digestion products

Metabolic engineering and thermodynamic characterization of an extracellular -glucosidase produced by Aspergillus niger

Production and partial purification of α-amylase from Pseudomonas sp. 2 under solid-state fermentation

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