Studies of effects of macromolecular crowding and confinement on protein folding and protein stability

2004 ◽  
Vol 17 (5) ◽  
pp. 433-440 ◽  
Author(s):  
Guanghui Ping ◽  
Jian-Min Yuan ◽  
Zhengfei Sun ◽  
Yen Wei
Keyword(s):  
Protein Folding ◽  
Protein Stability ◽  
Macromolecular Crowding

scholarly journals Differential strengths of molecular determinants guide environment specific mutational fates

2017 ◽  
Author(s):  
Rohan Dandage ◽  
Rajesh Pandey ◽  
Gopal Jayaraj ◽  
Kausik Chakraborty
Keyword(s):  
Protein Folding ◽  
Molecular Evolution ◽  
Protein Stability ◽  
Environmental Effects ◽  
Environmental Conditions ◽  
Fitness Landscape ◽  
Dependent Manner ◽  
The Common ◽  
Physical And Chemical ◽  
Substrate Binding Protein

AbstractUnder the influence of selection pressures imposed by natural environments, organisms maintain competitive fitness through underlying molecular evolution of individual genes across the genome. For molecular evolution, how multiple interdependent molecular constraints play a role in determination of fitness under different environmental conditions is largely unknown. Here, using Deep Mutational Scanning (DMS), we quantitated empirical fitness of ∼2000 single site mutants of Gentamicin-resistant gene (GmR). This enabled a systematic investigation of effects of different physical and chemical environments on the fitness landscape of the gene. Molecular constraints of the fitness landscapes seem to bear differential strengths in an environment dependent manner. Among them, conformity of the identified directionalities of the environmental selection pressures with known effects of the environments on protein folding proves that along with substrate binding, protein stability is the common strong constraint of the fitness landscape. Our study thus provides mechanistic insights into the molecular constraints that allow accessibility of mutational fates in environment dependent manner.Author SummaryEnvironmental conditions play a central role in both organismal adaptations and underlying molecular evolution. Understanding of environmental effects on evolution of genotype is still lacking a depth of mechanistic insights needed to assist much needed ability to forecast mutational fates. Here, we address this issue by culminating high throughput mutational scanning using deep sequencing. This approach allowed comprehensive mechanistic investigation of environmental effects on molecular evolution. We monitored effects of various physical and chemical environments onto single site mutants of model antibiotic resistant gene. Alongside, to get mechanistic understanding, we identified multiple molecular constraints which contribute to various degrees in determining the resulting survivabilities of mutants. Across all tested environments, we find that along with substrate binding, protein stability stands out as the common strong constraints. Remarkable direct dependence of the environmental fitness effects on the type of environmental alteration of protein folding further proves that protein stability is the major constraint of the gene. So, our findings reveal that under the influence of environmental conditions, mutational fates are channeled by various degrees of strengths of underlying molecular constraints.

scholarly journals Residue-Level Interrogation of Macromolecular Crowding Effects on Protein Stability

2008 ◽  
Vol 130 (21) ◽  
pp. 6826-6830 ◽  
Author(s):  
Lisa M. Charlton ◽  
Christopher O. Barnes ◽  
Conggang Li ◽  
Jillian Orans ◽  
Gregory B. Young ◽  
...  
Keyword(s):  
Protein Stability ◽  
Macromolecular Crowding ◽  
Residue Level ◽  
Crowding Effects

Effect of Dextran on Protein Stability and Conformation Attributed to Macromolecular Crowding

2003 ◽  
Vol 326 (4) ◽  
pp. 1227-1237 ◽  
Author(s):  
Kenji Sasahara ◽  
Peter McPhie ◽  
Allen P. Minton
Keyword(s):  
Protein Stability ◽  
Macromolecular Crowding

scholarly journals Oxidative folding of hirudin in human serum

2006 ◽  
Vol 394 (1) ◽  
pp. 249-257 ◽  
Author(s):  
Jui-Yoa Chang ◽  
Bao-Yun Lu ◽  
Por-Hsiung Lai
Keyword(s):  
Amino Acids ◽  
Protein Folding ◽  
Human Serum ◽  
Initial Phase ◽  
Macromolecular Crowding ◽  
Specific Inhibitor ◽  
Crowding Effect ◽  
Oxidative Folding ◽  
Disulphide Bonds ◽  
Kinetics Of

Human serum contains factors that promote oxidative folding of disulphide proteins. We demonstrate this here using hirudin as a model. Hirudin is a leech-derived thrombin-specific inhibitor containing 65 amino acids and three disulphide bonds. Oxidative folding of hirudin in human serum is shown to involve an initial phase of rapid disulphide formation (oxidation) to form the scrambled isomers as intermediates. This is followed by the stage of slow disulphide shuffling of scrambled isomers to attain the native hirudin. The kinetics of regenerating the native hirudin depend on the concentrations of both hirudin and human serum. Quantitative regeneration of native hirudin in undiluted human serum can be completed within 48 h, without any redox supplement. These results cannot be adequately explained by the existing oxidized thiol agents in human serum or the macromolecular crowding effect, and therefore indicate that human serum may contain yet to be identified potent oxidase(s) for assisting protein folding.

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