Production, purification and characterization of raw starch hydrolyzing thermostable acidic α-amylase from hot springs, India

Sudan, Sarabjeet Kour; Kumar, Narender; Kaur, Ishwinder; Sahni, Girish

doi:10.1016/j.ijbiomac.2018.05.231

Cited by 56 publications

(40 citation statements)

References 40 publications

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“…WBC is one of the largest subfamilies of ABC transporters, a superfamily of proteins found in many organisms ranging from prokaryotes to eukaryotes. ABC transporters contain two nucleotide binding domains and two trans‐membrane domains (Sudan, Kumar, Kaur, & Sahni, ). The Oswbc11 mutant had a base substitution on the last exon (C to T) which resulted in a premature stop codon within the trans‐membrane domain.…”

Section: Discussionmentioning

confidence: 99%

Identification and gene cloning of a male‐sterile mutant Oswbc11 in rice

Xuan

Liu

et al. 2019

Plant Breeding

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Commercial application of nuclear genetic male sterility has been improved recently by a novel technique, Seed Production Technology (SPT), which incorporates transgenic maintainer lines capable of propagating non‐transgenic nuclear male‐sterile lines for use as female parents in hybrid production. Here, we identified a rice nuclear male‐sterile mutant, Oswbc11, with abnormal pollen development and lipid transport. We finely mapped the Oswbc11 gene into a 12.5‐kb region on chromosome 10 and found one candidate gene, which had a base substitution (C to T) resulting in a premature stop codon and was functionally confirmed by CRISPR/Cas9 technology. Gene OsWBC11 encodes a sub‐family member of adenosine triphosphate‐binding cassette transporter, which participates in the active transport of a wide range of molecules across membranes. Moreover, the agronomic traits of Oswbc11 mutants showed no significant differences compared to the wild‐type control except for the seed setting rate. These results indicated that Oswbc11 gene could be used in rice hybrid breeding as a recessive nuclear male‐sterile gene combined with CRISPR/Cas9 and the SPT technology and applied in different rice varieties.

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Section: Discussionmentioning

confidence: 99%

Identification and gene cloning of a male‐sterile mutant Oswbc11 in rice

Xuan

Liu

et al. 2019

Plant Breeding

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“…Purification of α-amylase is often done by a combination of several methods. The initial step was mostly done by salt precipitation, followed by chromatograpy methods (Wu et al, 2018;Du et al, 2018;Sudan et al, 2018). Of the five fractions after ammonium sulfate precipitation, Fraction 2 produced the highest activity (42.4 U/ mL) and protein concentration (2.09 mg/mL).…”

Section: Ammonium Sulfat Precipitationmentioning

confidence: 99%

“…The concentration of ammonium sulfate needed to produce optimum α-amylase specific activity varies. Precipitation of α-amylase is commonly carried out at concentrations of 60-80% (Al-Quadan et al, 2011;Sudan et al, 2018), although there is an α-amylase precipitated in only 30% concentration of salt (Wu et al, 2018). The surface of proteins from thermophilic bacteria contains plenty of charged residues.…”

Section: Ammonium Sulfat Precipitationmentioning

confidence: 99%

“…Occasionally, a combination of chromatography methods is needed to increase the enzyme purity. For example, combination of exclusion gel (Sephadex G100) and ion exchange (DEAE Sepharose) chromatography increases the purity of about 30% (Al-Quadan et al, 2101), while the combination of Q Sepharose and Superdex 75PG increases the purity by eight folds (Sudan et al, 2018). This, again, may occur due to differences in the interactions of the purified protein with the substance used for purification, in this case, ammonium sulfate and column matrix.…”

Section: Evaluation Of the Purification Stepsmentioning

confidence: 99%

“…While the exploration of new microorganisms from extreme environments to produce more stable enzymes has been done for many years, studies to find new α-amylases continue being carried out (Krüger et al, 2018). Researches to obtain new α-amylase with better stability are often done by isolating extremophiles from various sources, such as hot spring (Sudan et al, 2018;Wu et al, 2018), deep sea (Jiang et al, 2015), and even honey (Du et al, 2018). Various studies have also been done to improve the stability of the enzyme through genetic engineering, immobilization, chemical modification and protein engineering (Dey et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

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Purification and Partial Characterization of α-Amylase Produced by a Thermo-Halophilic Bacterium Isolate PLS 75

2018

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Bio-based industries require stable enzymes in a broad range of environmental conditions. Extremophiles have attracted more interests as the source of such enzymes, one of which is α-amylase. This study aimed to purify and characterize α-amylase produced by a thermo-halophilic bacterium PLS 75 isolated from underwater fumaroles. Ammonium sulfate precipitation results showed that the highest specific α-amylase activity (21.7 U/mg) obtained at 40-60% saturation level, with a purity of 7.7-fold of the crude extract with 16.2% yield. Further purification using DEAE Sepharose column chromatography increased the enzyme purity 11.1-fold of the crude extract with 7.1% yield. Specific activity after column chromatography purification was 31.3 U/mg. The pure enzyme had a low molecular weight of 14 kDa. The enzyme showed the highest activity at 80 °C and pH 5. The activity increased to 126% when in methanol, while decreased when in ethyl acetate and chloroform. The characteristics of α-amylase with low molecular weight, which was active in acidic condition, stable in polar and non-polar solvents, may be used for for specific industrial needs.

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Purification and Characterization of an Amylase from a Newly Isolated Geobacillus thermoleovorans TTIO4 from the Hot Spring of Tuwa‐Timba

Shukla

Singh

2023

Starch Stärke

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A thermophilic amylase produced by a newly isolated Geobacillus thermoleovorans TTIO4 from a hot spring in India is investigated. The isolate TTIO4 optimally produces extracellular amylase at 55 °C, pH 7, and 24 h of growth. The amylase is purified by 42 fold with a yield of 9.7%. The enzyme has a molecular weight of 40 kDa. The purified amylase catalyzes the reaction optimally at 60 °C and pH 7. Besides, it displays maximum thermal stability at 50 °C for 18 h. While the catalyst is improved by Ca+2, it is wholly inhibited by Hg+2. Furthermore, the chelators, inhibitors and solvents adversely affect the enzyme activity. The amylase is stabilized by SDS, while other surfactants negatively affect the enzyme. Km, Vmax, Kcat, and Kcat/Km of the amylase are 0.26 mg mL−1, 192.3 µg mL−1 min−1, 1.92 s−1, and 7.383 s−1 mg−1, respectively. The deactivation constant, half‐life, and change in Gibb's free energy, enthalpy, and entropy are determined at 50–70 °C, which supports the thermal stability and substrate affinity of the amylase. The washing quality of commercial detergents is distinctly enhanced when supplemented with this enzyme. Therefore, this amylase possesses significant thermal and surfactant stability has significant commercial potential.

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Production, purification and characterization of raw starch hydrolyzing thermostable acidic α-amylase from hot springs, India

Cited by 56 publications

References 40 publications

Identification and gene cloning of a male‐sterile mutant Oswbc11 in rice

Identification and gene cloning of a male‐sterile mutant Oswbc11 in rice

Purification and Partial Characterization of α-Amylase Produced by a Thermo-Halophilic Bacterium Isolate PLS 75

Purification and Characterization of an Amylase from a Newly Isolated Geobacillus thermoleovorans TTIO4 from the Hot Spring of Tuwa‐Timba

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