The role of proline substitutions within flexible regions on thermostability of luciferase

Yu, Haoran; Zhao, Yang; Guo, Chao; Gan, Yiru; Huang, He

doi:10.1016/j.bbapap.2014.10.017

Cited by 84 publications

(50 citation statements)

References 45 publications

(45 reference statements)

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“…The DCl-of Met system is larger than that of Pro system, which indicates that the migration speed of Cl -ion Met inhibition films is slower than that in Pro inhibition films. This may be because the molecular shape of Pro molecule is smaller and the molecular rigidity is stronger than Met molecule, thus, the compactness or quality of Pro inhibition films is higher than that of Met inhibition films [44][45][46] . Even though, this molecular structure advantages are not reflected on the single Met and Pro inhibitors due to the adsorption capability of Met molecule is stronger than that of Pro molecule on iron surface, the inhibition effect of Met/Pro compound inhibitor can be enhanced by Met molecule combining with Pro molecule due to this molecular structure advantages.…”

Section: Dynamic Simulationmentioning

confidence: 90%

“…The decreasing in the Cdl value is normally attributed to the replacement of the adsorbed water molecules at the metal surface by Met molecules that have a lower dielectric constant [43] . Thus, it can be concluded that the Met molecule have adsorbed on steel surface, then formed a homogeneous film, and the adsorption number perfect due to the molecular shape of Pro molecule is smaller and the molecular rigidity is stronger than Met molecule [44][45][46] . When the concentration of Pro is high, the quality of Met/Pro inhibition films is poor due to the competitive adsorption between Met and Pro molecules on iron surface.…”

Section: Molecular Dynamic Simulationsmentioning

confidence: 96%

“…Thus, while the concentration of Pro is low, the Met and Pro molecules can adsorb on iron surface and 11 form a dense film with a major component of Met molecule and a minor component of the Pro molecule. Pro molecule can fill in the leak of Met inhibition films and make the films perfect due to the molecular shape of Pro molecule is smaller and the molecular rigidity is stronger than Met molecule [44][45][46] . When the concentration of Pro is high, the quality of Met/Pro inhibition films is poor due to the competitive adsorption between Met and Pro molecules on iron surface.…”

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confidence: 98%

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Inhibition of carbon steel corrosion in phase-change-materials solution by methionine and proline

et al. 2016

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Section: Dynamic Simulationmentioning

confidence: 90%

Section: Molecular Dynamic Simulationsmentioning

confidence: 96%

mentioning

confidence: 98%

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Inhibition of carbon steel corrosion in phase-change-materials solution by methionine and proline

et al. 2016

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“…One approach is based on engineering of flexible residues by rigidifying them through RMSF analysis [39]. One critical point should be considered is, this manipulation should not affect the required flexibility for enzymatic reaction.…”

Section: Discussionmentioning

confidence: 99%

Structural and dynamical insight into thermally induced functional inactivation of firefly luciferase

2017

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Luciferase is the key component of light production in bioluminescence process. Extensive and advantageous application of this enzyme in biotechnology is restricted due to its low thermal stability. Here we report the effect of heating up above Tm on the structure and dynamical properties of luciferase enzyme compared to temperature at 298 K. In this way we demonstrate that the number of hydrogen bonds between N- and C-domain is increased for the free enzyme at 325 K. Increased inter domain hydrogen bonds by three at 325 K suggests that inter domain contact is strengthened. The appearance of simultaneous strong salt bridge and hydrogen bond between K529 and D422 and increased existence probability between R533 and E389 could mechanistically explain stronger contact between N- and C-domain. Mutagenesis studies demonstrated the importance of K529 and D422 experimentally. Also the significant reduction in SASA for experimentally important residues K529, D422 and T343 which are involved in active site region was observed. Principle component analysis (PCA) in our study shows that the dynamical behavior of the enzyme is changed upon heating up which mainly originated from the change of motion modes and associated extent of those motions with respect to 298 K. These findings could explain why heating up of the enzyme or thermal fluctuation of protein conformation reduces luciferase activity in course of time as a possible mechanism of thermal functional inactivation. According to these results we proposed two strategies to improve thermal stability of functional luciferase.

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“…The selection of mutation sites in enzymes or proteins is the most critical step in the rational design method [36]. Inappropriate positions can affect folding and destroy the structure of an active protein [37]. Therefore, two main selection principles have been considered: one was that the chosen mutation sites should be situated away from the active center to guarantee that the overall structure and catalytic activity of the target enzyme were less affected.…”

Section: Discussionmentioning

confidence: 99%

A Novel Strategy for Thermostability Improvement of Trypsin Based on N-Glycosylation within the ��-Loop Region

Guo¹,

Liu²,

Yu³

et al. 2016

Journal of Microbiology and Biotechnology

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The Ω-loop is a nonregular and flexible structure that plays an important role in molecular recognition, protein folding, and thermostability. In the present study, molecular dynamics simulation was carried out to assess the molecular stability and flexibility profile of the porcine trypsin structures. Two Ω-Loops (fragment 57-67 and fragment 78-91) were confirmed to represent the flexible region. Subsequently, glycosylation site-directed mutations (A73S, N84S, and R104S) were introduced within the Ω-loop region and its wing chain based on its potential N-glycosylation sites (Asn-Xaa-Ser/Thr consensus sequences) and structure information to improve the thermostability of trypsin. The result demonstrated that the halflife of the N84S mutant at 50°C increased by 177.89 min when compared with that of the wildtype enzyme. Furthermore, the significant increase in the thermal stability of the N84S mutant has also been proven by an increase in the Tm values determined by circular dichroism. Additionally, the optimum temperatures of the wild-type enzyme and the N84S mutant were 75°C and 80°C, respectively. In conclusion, we obtained the thermostability-improved enzyme N84S mutant, and the strategy used to design this mutant based on its structural information and N-linked glycosylation modification could be applied to engineer other enzymes to meet the needs of the biotechnological industry.

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The role of proline substitutions within flexible regions on thermostability of luciferase

Cited by 84 publications

References 45 publications

Inhibition of carbon steel corrosion in phase-change-materials solution by methionine and proline

Inhibition of carbon steel corrosion in phase-change-materials solution by methionine and proline

Structural and dynamical insight into thermally induced functional inactivation of firefly luciferase

A Novel Strategy for Thermostability Improvement of Trypsin Based on N-Glycosylation within the ��-Loop Region

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