Insights into the unfolding pathway and identification of thermally sensitive regions of phytase from Aspergillus niger by molecular dynamics simulations

Kumar, Kranti; Patel, Kamlesh; Agrawal, Dinesh Chandra; Khire, J. M.

doi:10.1007/s00894-015-2696-z

Cited by 8 publications

(6 citation statements)

References 54 publications

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“…The number of hydrogen, ionic and disulfide bond interactions is believed to have a strong correlation with thermostability (Zhang 2007;Liao, et al, 2013;Fei et al, 2013;Hesampour et al, 2015;Kumar et al, 2015;Tan et al, 2016;Han et al, 2018;Li et al, 2019;Zhang et al, 2020). However, here we show that, for these enzymes, this is not true.…”

Section: Quantification and Analysis Of Interactions Of The Structura...contrasting

confidence: 50%

“…The association between the structure and the biochemical properties of enzymes can provide relevant information about factors that affect stability and catalytic activity. Several methods have been used for this purpose, including comparative sequence, structure analysis, molecular dynamics studies, or even the evaluation of random or rational mutations on enzyme activity (Zhang 2007;Liao, et al, 2013;Fei et al, 2013;Hesampour et al, 2015;Kumar et al, 2015;Tan et al, 2016;Han et al, 2018;Li et al, 2019;Zhang et al, 2020;Acquistapace et al, 2022). In this study, we report a comparative approach between sequences, threedimensional structures and biochemical characteristics of phytases already characterized and described in the literature, adopting factors such as: hydrogen, ionic and Van der Waals (VdW) interactions and disulfide bonds.…”

Section: Introductionmentioning

confidence: 99%

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Sequence diversity and catalytic properties of phytases

Pires

Polêto

Vidigal

et al. 2022

RSD

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Phytic acid is an antinutritional factor in cereal feeds, and the use of phytases increases the bioavailability of nutrients bound to this molecule. However, the application of these enzymes depends on their thermal stability and activity at acidic pH. Therefore, in this study we created a database composed of 59 phytase sequences and analyzed the interactions that stabilize their structures in order to understand whether they contribute to the biochemical properties observed. The sequences were aligned and grouped at 30 % similarity, generating 5 clusters, which highlights the high variability among them. A comparative structural analysis of the cluster 3 phytases revealed conserved catalytic domains, as well as eight cysteine residues along the primary sequence, forming disulfide bonds for stabilizing the three-dimensional structure. However, the number of Van der Waals, ionic, and hydrogen interactions, and disulfide bonds was not determinant for the biochemical characteristics presented by these enzymes. The phytase KM873028, from cluster 3, was selected for characterization studies, but its expression in Pichia pastoris generated a protein with properties distinct from those derived from the same sequence expressed in a prokaryotic system. It is likely that the differences observed are associated with the location of the interactions in the structures, non-conserved amino acid residues found around the catalytic site, and post-translational modifications inherent to the expression systems. These possibilities highlight the relevance of strategic choices related to enzyme expression aiming at its production and industrial feasibility.

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Section: Quantification and Analysis Of Interactions Of The Structura...contrasting

confidence: 50%

Section: Introductionmentioning

confidence: 99%

Sequence diversity and catalytic properties of phytases

Pires

Polêto

Vidigal

et al. 2022

RSD

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show abstract

“…Some regions of PHYA from A. niger (the salt bridges between Glu125-Arg163, Asp299-Arg136, Asp266-Arg219, Asp339-Lys278, Asp335-Arg136, and Asp424-Arg428) were predicted as thermolabile by bioinformatic tools given their expansion upon the increase of temperature. This data could be the basis to engineer novel thermostable phytases with desirable properties [102].…”

Section: Thermostability: An Issue For Industrial Applicationmentioning

confidence: 96%

Fungal phytases: from genes to applications

Corrêa

Araújo

2020

Braz J Microbiol

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Phytic acid stores 60-90% of the inorganic phosphorus in legumes, oil seeds, and cereals, making it inaccessible for metabolic processes in living systems. In addition, given its negative charge, phytic acid complexes with divalent cations, starch, and proteins. Inorganic phosphorous can be released from phytic acid upon the action of phytases. Phytases are phosphatases produced by animals, plants, and microorganisms, notably Aspergillus niger, and are employed as animal feed additive, in chemical industry and for ethanol production. Given the industrial relevance of phytases produced by filamentous fungi, this work discusses the functional characterization of fungal phytase-coding genes/proteins, highlighting the physicochemical parameters that govern the enzymatic activity, the development of phytase super-producing strains, and key features for industrial applications.

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“…By increasing the temperature, most thermally sensitive regions, including loops 8, 10, 14, 17 and 24, showed intense increments in RMSF values. It was concluded that the location of the salt bridge was more important compared to the number of salt bridges for thermostability [22].…”

Section: Aspergillus Phytasesmentioning

confidence: 99%

Integrative Structural and Computational Biology of Phytases for the Animal Feed Industry

et al. 2020

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Resistance to high temperature, acidic pH and proteolytic degradation during the pelleting process and in the digestive tract are important features of phytases as animal feed. The integration of insights from structural and in silico analyses into factors affecting thermostability, acid stability, proteolytic stability, catalytic efficiency and specific activity, as well as N-glycosylation, could improve the limitations of marginal stable biocatalysts with trade-offs between stability and activity. Synergistic mutations give additional benefits to single substitutions. Rigidifying the flexible loops or inter-molecular interactions by reinforcing non-bonded interactions or disulfide bonds, based on structural and roof mean square fluctuation (RMSF) analyses, are contributing factors to thermostability. Acid stability is normally achieved by targeting the vicinity residue at the active site or at the neighboring active site loop or the pocket edge adjacent to the active site. Extending the positively charged surface, altering protease cleavage sites and reducing the affinity of protease towards phytase are among the reported contributing factors to improving proteolytic stability. Remodeling the active site and removing steric hindrance could enhance phytase activity. N-glycosylation conferred improved thermostability, proteases degradation and pH activity. Hence, the integration of structural and computational biology paves the way to phytase tailoring to overcome the limitations of marginally stable phytases to be used in animal feeds.

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Insights into the unfolding pathway and identification of thermally sensitive regions of phytase from Aspergillus niger by molecular dynamics simulations

Cited by 8 publications

References 54 publications

Sequence diversity and catalytic properties of phytases

Sequence diversity and catalytic properties of phytases

Fungal phytases: from genes to applications

Integrative Structural and Computational Biology of Phytases for the Animal Feed Industry

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