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Optimization to low temperature activity in psychrophilic enzymes.

Struvay C, Feller G - Int J Mol Sci (2012)

Bottom Line: Most psychrophilic enzymes optimize a high activity at low temperature at the expense of substrate affinity, therefore reducing the free energy barrier of the transition state.In these naturally evolved enzymes, the optimization to low temperature activity is reached via destabilization of the structures bearing the active site or by destabilization of the whole molecule.This involves a reduction in the number and strength of all types of weak interactions or the disappearance of stability factors, resulting in improved dynamics of active site residues in the cold.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biochemistry, Centre for Protein Engineering, University of Liège, Institute of Chemistry B6a, B-4000 Liège-Sart Tilman, Belgium; E-Mail: cstruvay@ulg.ac.be.

ABSTRACT
Psychrophiles, i.e., organisms thriving permanently at near-zero temperatures, synthesize cold-active enzymes to sustain their cell cycle. These enzymes are already used in many biotechnological applications requiring high activity at mild temperatures or fast heat-inactivation rate. Most psychrophilic enzymes optimize a high activity at low temperature at the expense of substrate affinity, therefore reducing the free energy barrier of the transition state. Furthermore, a weak temperature dependence of activity ensures moderate reduction of the catalytic activity in the cold. In these naturally evolved enzymes, the optimization to low temperature activity is reached via destabilization of the structures bearing the active site or by destabilization of the whole molecule. This involves a reduction in the number and strength of all types of weak interactions or the disappearance of stability factors, resulting in improved dynamics of active site residues in the cold. Considering the subtle structural adjustments required for low temperature activity, directed evolution appears to be the most suitable methodology to engineer cold activity in biological catalysts.

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Related in: MedlinePlus

Inactivation and unfolding of psychrophilic enzymes. The activity of psychrophilic enzymes (upper panel, blue line) is inactivated by temperature before unfolding of the protein structure (lower panel, blue line) illustrating the pronounced heat-lability of the active site. By contrast, inactivation of mesophilic enzymes (black curves) closely corresponds to the loss of the protein conformation. Adapted from [14].
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f2-ijms-13-11643: Inactivation and unfolding of psychrophilic enzymes. The activity of psychrophilic enzymes (upper panel, blue line) is inactivated by temperature before unfolding of the protein structure (lower panel, blue line) illustrating the pronounced heat-lability of the active site. By contrast, inactivation of mesophilic enzymes (black curves) closely corresponds to the loss of the protein conformation. Adapted from [14].

Mentions: Most psychrophilic enzymes share at least one property: a heat-labile activity, irrespective of the protein structural stability. Furthermore, the active site appears to be the most heat-labile structural element of these proteins [13–15]. Figure 2 illustrates this significant difference between the stability of the active site and the stability of the structure. The lower panel shows the stability of the structure as recorded by fluorescence. As expected, the structure of the cold-active enzyme is less stable than the mesophilic one. In the upper panel, the activity is recorded under the same experimental conditions and it can be seen that the mesophilic enzyme is inactivated when the protein unfolds. By contrast, activity of the cold-active enzyme is lost before the protein unfolds. This means that the active site is even more heat-labile than the whole protein structure. It was also shown that the active site of a psychrophilic -amylase is the first structural element that unfolds in transverse urea gradient gel electrophoresis [16]. All these aspects point to a very unstable and flexible active site and illustrate a central concept in cold adaptation: localized increases in flexibility at the active site are responsible for the high but heat-labile activity [17], whereas other regions of the enzyme might or might not be characterized by low stability when not involved in catalysis. For instance, psychrophilic carbonic anhydrase [18] and isocitrate dehydrogenase [19] are highly stable enzymes with however improved flexibility in regions driving catalysis.


Optimization to low temperature activity in psychrophilic enzymes.

Struvay C, Feller G - Int J Mol Sci (2012)

Inactivation and unfolding of psychrophilic enzymes. The activity of psychrophilic enzymes (upper panel, blue line) is inactivated by temperature before unfolding of the protein structure (lower panel, blue line) illustrating the pronounced heat-lability of the active site. By contrast, inactivation of mesophilic enzymes (black curves) closely corresponds to the loss of the protein conformation. Adapted from [14].
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3472767&req=5

f2-ijms-13-11643: Inactivation and unfolding of psychrophilic enzymes. The activity of psychrophilic enzymes (upper panel, blue line) is inactivated by temperature before unfolding of the protein structure (lower panel, blue line) illustrating the pronounced heat-lability of the active site. By contrast, inactivation of mesophilic enzymes (black curves) closely corresponds to the loss of the protein conformation. Adapted from [14].
Mentions: Most psychrophilic enzymes share at least one property: a heat-labile activity, irrespective of the protein structural stability. Furthermore, the active site appears to be the most heat-labile structural element of these proteins [13–15]. Figure 2 illustrates this significant difference between the stability of the active site and the stability of the structure. The lower panel shows the stability of the structure as recorded by fluorescence. As expected, the structure of the cold-active enzyme is less stable than the mesophilic one. In the upper panel, the activity is recorded under the same experimental conditions and it can be seen that the mesophilic enzyme is inactivated when the protein unfolds. By contrast, activity of the cold-active enzyme is lost before the protein unfolds. This means that the active site is even more heat-labile than the whole protein structure. It was also shown that the active site of a psychrophilic -amylase is the first structural element that unfolds in transverse urea gradient gel electrophoresis [16]. All these aspects point to a very unstable and flexible active site and illustrate a central concept in cold adaptation: localized increases in flexibility at the active site are responsible for the high but heat-labile activity [17], whereas other regions of the enzyme might or might not be characterized by low stability when not involved in catalysis. For instance, psychrophilic carbonic anhydrase [18] and isocitrate dehydrogenase [19] are highly stable enzymes with however improved flexibility in regions driving catalysis.

Bottom Line: Most psychrophilic enzymes optimize a high activity at low temperature at the expense of substrate affinity, therefore reducing the free energy barrier of the transition state.In these naturally evolved enzymes, the optimization to low temperature activity is reached via destabilization of the structures bearing the active site or by destabilization of the whole molecule.This involves a reduction in the number and strength of all types of weak interactions or the disappearance of stability factors, resulting in improved dynamics of active site residues in the cold.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biochemistry, Centre for Protein Engineering, University of Liège, Institute of Chemistry B6a, B-4000 Liège-Sart Tilman, Belgium; E-Mail: cstruvay@ulg.ac.be.

ABSTRACT
Psychrophiles, i.e., organisms thriving permanently at near-zero temperatures, synthesize cold-active enzymes to sustain their cell cycle. These enzymes are already used in many biotechnological applications requiring high activity at mild temperatures or fast heat-inactivation rate. Most psychrophilic enzymes optimize a high activity at low temperature at the expense of substrate affinity, therefore reducing the free energy barrier of the transition state. Furthermore, a weak temperature dependence of activity ensures moderate reduction of the catalytic activity in the cold. In these naturally evolved enzymes, the optimization to low temperature activity is reached via destabilization of the structures bearing the active site or by destabilization of the whole molecule. This involves a reduction in the number and strength of all types of weak interactions or the disappearance of stability factors, resulting in improved dynamics of active site residues in the cold. Considering the subtle structural adjustments required for low temperature activity, directed evolution appears to be the most suitable methodology to engineer cold activity in biological catalysts.

Show MeSH
Related in: MedlinePlus