Probing the role of asparagine mutation in thermostability of Bacillus KR-8104 α-amylase

Rahimzadeh, Mahsa; Khajeh, Khosro; Mirshahi, Manoochehr; Khayatian, Mahmood; Schwarzenbacher, Robert

doi:10.1016/j.ijbiomac.2011.11.014

Cited by 23 publications

(14 citation statements)

References 46 publications

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“…The method that has been used to analyze the heat inactivation rate constants of L-asparaginase requires enzyme incubation at desirable temperatures and cooling in ice before measurement of the residual activity [25, 46]. After incubation at 37˚C and 40˚C, the half-lives (t 1/2 ) of Bacillus PG03 and Bacillus PG04 were measured (Table 1 ).…”

Section: Resultsmentioning

confidence: 99%

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Purification, Characterization and Comparison between Two New L-asparaginases from PG03 and PG04

Rahimzadeh¹,

Poodat²,

Javadpour³

et al. 2016

Open Biochem J.

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Background:L-asparaginase has been used as a chemotherapeutic agent in treatment of lymphoblastic leukemia. In the present investigation, Bacillus sp. PG03 and Bacillus sp. PG04 were studied.Methods:L- asparaginases were produced using different culture media and were purified using ion exchange chromatography.Results:Maximum productivity was obtained when asparagine was used as the nitrogen source at pH 7 and 48 h after cultivation. New intracellular L-asparaginases showed an apparent molecular weight of 25 kDa and 30 kDa by SDS-PAGE respectively. These enzymes were active in a wide pH range (3-9) with maximum activity at pH 6 for Bacillus PG03 and pH 7 for Bacillus PG04 L-asparaginase. Bacillus PG03 enzyme was optimally active at 37 ˚C and Bacillus PG04 maximum activity was observed at 40˚C. Kinetic parameters km and Vmax of both enzymes were studied using L-asparagine as the substrate. Thermal inactivation studies of Bacillus PG03 and Bacillus PG04 L-asparaginase exhibited t1/2 of 69.3 min and 34.6 min in 37 ˚C respectively. Also T50 and ∆G of inactivation were measured for both enzymes.Conclusion: The results revealed that both enzymes had appropriate characteristics and thus could be a potential candidate for medical applications.

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Section: Resultsmentioning

confidence: 99%

“…Plots of the log of residual activity versus time were linear, indicating a first-order decay. T 50 is the temperature of incubation at which 50% of the initial enzyme activity is lost during 30 min incubation [25]. The ΔG* parameter of the L-asparaginase enzyme was determined as follows [26]:…”

Section: Methodsmentioning

confidence: 99%

Purification, Characterization and Comparison between Two New L-asparaginases from PG03 and PG04

Rahimzadeh¹,

Poodat²,

Javadpour³

et al. 2016

Open Biochem J.

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“…In this sub-cluster three thermo-stable α-amylases (EC # 3.2.1.1) mutants from the genus Bacillus can be found. In two of the three cases (3DC0, Rahimzadeh et al, 2012;1BF2, Fujimoto et al, 1998) the directed mutagenesis was performed to increase thermal stability. In the case of 1UA7, it is a mutant of the catalytic site that is not supposed to change stability or function with respect to the wild type [Kagawa et al, 2003].…”

Section: Npc1 Dataset: Analysis Of the Residue Variation In The Contementioning

confidence: 99%

Protein structures as shapes: Analysing protein structure variation using geometric morphometrics

Hleap

Blouin

2017

Preprint

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A phenotype is defined as an organism's physical traits. In the macroscopic world, an animal's shape is a phenotype. Geometric morphometrics (GM) can be used to analyze its shape. Let's pose protein structures as microscopic three dimensional shapes, and apply principles of GM to the analysis of macromolecules. In this paper we introduce a way to 1) abstract a structure as a shape; 2) align the shapes; and 3) perform statistical analysis to establish patterns of variation in the datasets. We show that general procrustes superimposition (GPS) can be replaced by multiple structure alignment without changing the outcome of the test. We also show that estimating the deformation of the shape (structure) can be informative to analyze relative residue variations. Finally, we show an application of GM for two protein structure datasets: 1) in the α-amylase dataset we demonstrate the relationship between structure, function, and how the 1 . CC-BY-NC-ND 4.0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/219030 doi: bioRxiv preprint first posted online Nov. 13, 2017; dependency of chloride has an important effect on the structure; and 2) in the Niemann-Pick disease, type C1 (NPC1) protein's molecular dynamic simulation dataset, we introduce a simple way to analyze the trajectory of the simulation by means of protein structure variation.

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“…The plots were linear indicating first-order decay. Another parameter which gives information about thermal stability was T 50 which is the temperature of incubation at which 50% of the initial enzyme activity is lost during 30 min incubation (Rahimzadeh et al, 2012). Temperatures from 30 to 50°C were used for T 50 measurements.…”

Section: Thermal Stability Assaymentioning

confidence: 99%

Intracellular L-Asparaginase from <i>Bacillus</i> sp.PG02: Purification, Biochemical Characterization and Evaluation of Optimum pH and Temperature

Qeshmi

Rahimzadeh

Javadpour

et al. 2016

American Journal of Biochemistry and Biotechnology

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Bacterial L-asparaginases are amidohydrolases that act on Lasparagine and produce L-aspartate and ammonia. These enzymes have been used in treatment of lymphoblastic leukemia. In the present study, a novel strain, Bacillus sp. PG02 was explored for the production of intracellular L-asparaginase enzyme. The nitrogen source for L-asparaginase production was L-asparagine. New intracellular L-asparaginase was purified using ion exchange chromatography and the purity was assessed using SDS-PAGE. Kinetic parameters k m and V max and thermal properties were studied using L-asparagine as the substrate. SDS-PAGE analysis showed apparent molecular weight of approximately 38 kDa. The enzyme was active in a wide pH ranges (5-10) and it was maximally active at pH 7.5. Bacillus PG02 L-asparaginase was optimally active at 40°C. Thermal inactivation studies exhibited t 1/2 of 32.5 min in 37°C. Also T 50 and ∆G of inactivation were measured. The results revealed that the enzyme had appropriate characteristics and thus could be a potential candidate for medical and basic investigations.

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Probing the role of asparagine mutation in thermostability of Bacillus KR-8104 α-amylase

Cited by 23 publications

References 46 publications

Purification, Characterization and Comparison between Two New L-asparaginases from PG03 and PG04

Purification, Characterization and Comparison between Two New L-asparaginases from PG03 and PG04

Protein structures as shapes: Analysing protein structure variation using geometric morphometrics

Intracellular L-Asparaginase from <i>Bacillus</i> sp.PG02: Purification, Biochemical Characterization and Evaluation of Optimum pH and Temperature

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