A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake

Background: Cyclodextrin glucanotransferase (CGTase) is one of the most industrially important enzymes used in the commercial production of cyclodextrins (CDs). Alkaliphilic bacteria have attracted much interest in the last few decades because of their ability to produce extracellular enzymes that a...

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Autores principales:	Al-Sharawi,Samar Z.R., Ibrahim,Abdelnasser S.S., El-Shatoury,Einas H, Gebreel,Hassan M, Eldiwany,Ahmad
Lenguaje:	English
Publicado:	Pontificia Universidad Católica de Valparaíso 2013
Materias:	Amphibacillus sp cyclodextrin glycosyltransferase cyclodextrins Wadi Natrun soda lakes
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582013000600003
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spelling	oai:scielo:S0717-345820130006000032014-01-16A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lakeAl-Sharawi,Samar Z.R.Ibrahim,Abdelnasser S.S.El-Shatoury,Einas HGebreel,Hassan MEldiwany,Ahmad Amphibacillus sp cyclodextrin glycosyltransferase cyclodextrins Wadi Natrun soda lakes Background: Cyclodextrin glucanotransferase (CGTase) is one of the most industrially important enzymes used in the commercial production of cyclodextrins (CDs). Alkaliphilic bacteria have attracted much interest in the last few decades because of their ability to produce extracellular enzymes that are active and stable at high pH values. Here, we report the isolation of a new CGTase from alkaliphilic bacteria collected from Egyptian soda lakes and describe the purification and biochemical characterization of this CGTase. Results: Screening for CGTase-producing alkaliphilic bacteria from sediment and water samples collected from Egyptian soda lakes located in the Wadi Natrun valley resulted in the isolation of a potent CGTase-producing alkaliphilic bacterial strain, designated NRC-WN. Strain NRC-WN was belonging to genus Amplibacullus by 16S rDNA sequence analysis (similarity: ca. 98%). Among the tested nitrogen and carbon sources, peptone (0.15%, w/v) and soluble starch (0.4%, w/v) allowed maximal CGTase production by Amphibacillus sp. NRC-WN. CGTase was successfully purified from Amphibacillus sp. NRC-WN up to 159.7-fold through a combination of starch adsorption and anion exchange chromatography, resulting in a yield of 84.7%. SDS-PAGE analysis indicated that the enzyme was purified to homogeneity and revealed an estimated molecular mass of 36 kDa, which makes it one of the smallest CGTases reported in the literature. The purified enzyme exhibited maximum activity at 50ºC and was stable up to 70ºC, retaining 93% of its initial activity after treatment for 1 hr. Furthermore, Ca2+ ions (10 mM) significantly enhanced the thermal stability of the CGTase. The purified enzyme was active and stable over a wide pH range, showing maximal activity at pH 9.5. The enzyme was significantly stimulated by Zn2+, Ca2+ and Co2+ but was completely inhibited in the presence of Fe3+ and mercaptoethanol. The Km and Vmax values of the purified CGTase were estimated to be 0.0434 mg/ml and 3,333.3 mg β-CD/ml/min, respectively. β-CD was the predominant product of starch degradation by the Amphibacillus sp. NRC-WN CGTase, followed by α-and γ-CDs. Conclusions: A new low molecular mass alkaline CGTase was purified from a newly identified alkaliphilic Amphibacillus sp. NRC-WN isolate from the Egyptian soda lakes. The enzyme showed promising thermal and pH stability and a high affinity toward starch as a natural substrate.info:eu-repo/semantics/openAccessPontificia Universidad Católica de ValparaísoElectronic Journal of Biotechnology v.16 n.6 20132013-11-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582013000600003en10.2225/vol16-issue6-fulltext-18
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	Amphibacillus sp cyclodextrin glycosyltransferase cyclodextrins Wadi Natrun soda lakes
spellingShingle	Amphibacillus sp cyclodextrin glycosyltransferase cyclodextrins Wadi Natrun soda lakes Al-Sharawi,Samar Z.R. Ibrahim,Abdelnasser S.S. El-Shatoury,Einas H Gebreel,Hassan M Eldiwany,Ahmad A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake
description	Background: Cyclodextrin glucanotransferase (CGTase) is one of the most industrially important enzymes used in the commercial production of cyclodextrins (CDs). Alkaliphilic bacteria have attracted much interest in the last few decades because of their ability to produce extracellular enzymes that are active and stable at high pH values. Here, we report the isolation of a new CGTase from alkaliphilic bacteria collected from Egyptian soda lakes and describe the purification and biochemical characterization of this CGTase. Results: Screening for CGTase-producing alkaliphilic bacteria from sediment and water samples collected from Egyptian soda lakes located in the Wadi Natrun valley resulted in the isolation of a potent CGTase-producing alkaliphilic bacterial strain, designated NRC-WN. Strain NRC-WN was belonging to genus Amplibacullus by 16S rDNA sequence analysis (similarity: ca. 98%). Among the tested nitrogen and carbon sources, peptone (0.15%, w/v) and soluble starch (0.4%, w/v) allowed maximal CGTase production by Amphibacillus sp. NRC-WN. CGTase was successfully purified from Amphibacillus sp. NRC-WN up to 159.7-fold through a combination of starch adsorption and anion exchange chromatography, resulting in a yield of 84.7%. SDS-PAGE analysis indicated that the enzyme was purified to homogeneity and revealed an estimated molecular mass of 36 kDa, which makes it one of the smallest CGTases reported in the literature. The purified enzyme exhibited maximum activity at 50ºC and was stable up to 70ºC, retaining 93% of its initial activity after treatment for 1 hr. Furthermore, Ca2+ ions (10 mM) significantly enhanced the thermal stability of the CGTase. The purified enzyme was active and stable over a wide pH range, showing maximal activity at pH 9.5. The enzyme was significantly stimulated by Zn2+, Ca2+ and Co2+ but was completely inhibited in the presence of Fe3+ and mercaptoethanol. The Km and Vmax values of the purified CGTase were estimated to be 0.0434 mg/ml and 3,333.3 mg β-CD/ml/min, respectively. β-CD was the predominant product of starch degradation by the Amphibacillus sp. NRC-WN CGTase, followed by α-and γ-CDs. Conclusions: A new low molecular mass alkaline CGTase was purified from a newly identified alkaliphilic Amphibacillus sp. NRC-WN isolate from the Egyptian soda lakes. The enzyme showed promising thermal and pH stability and a high affinity toward starch as a natural substrate.
author	Al-Sharawi,Samar Z.R. Ibrahim,Abdelnasser S.S. El-Shatoury,Einas H Gebreel,Hassan M Eldiwany,Ahmad
author_facet	Al-Sharawi,Samar Z.R. Ibrahim,Abdelnasser S.S. El-Shatoury,Einas H Gebreel,Hassan M Eldiwany,Ahmad
author_sort	Al-Sharawi,Samar Z.R.
title	A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake
title_short	A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake
title_full	A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake
title_fullStr	A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake
title_full_unstemmed	A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake
title_sort	new low molecular mass alkaline cyclodextrin glucanotransferase from amphibacillus sp. nrc-wn isolated from an egyptian soda lake
publisher	Pontificia Universidad Católica de Valparaíso
publishDate	2013
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582013000600003
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A new low molecular mass alkaline cyclodextrin glucanotransferase from Amphibacillus sp. NRC-WN isolated from an Egyptian soda lake

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