The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases.
We hypothesize that aggregation of thermoalkalophilic lipases could be a thermostability mechanism. The conserved tryptophans (W211, W234) in the lid are of particular interest owing to their previous involvements in aggregation and thermostability mechanisms in many other proteins. The thermoalkalo...
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2013-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3877348?pdf=render |
| _version_ | 1818359861842280448 |
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| author | Emel Timucin O Ugur Sezerman |
| author_facet | Emel Timucin O Ugur Sezerman |
| author_sort | Emel Timucin |
| collection | DOAJ |
| description | We hypothesize that aggregation of thermoalkalophilic lipases could be a thermostability mechanism. The conserved tryptophans (W211, W234) in the lid are of particular interest owing to their previous involvements in aggregation and thermostability mechanisms in many other proteins. The thermoalkalophilic lipase from Bacillus thermocatenulatus (BTL2) and its mutants (W211A, W234A) were expressed and purified to homogeneity. We found that, when aggregated, BTL2 is more thermostable than its non-aggregating form, showing that aggregation potentiates thermostability in the thermoalkalophilic lipase. Among the two lid mutants, the W211A lowered aggregation tendency drastically and resulted in a much less thermostable variant of BTL2, which indicated that W211 stabilizes the intermolecular interactions in BTL2 aggregates. Further thermoactivity and CD spectroscopy analyses showed that W211A also led to a strong decrease in the optimal and the melting temperature of BTL2, implying stabilization by W211 also to the intramolecular interactions. The other lid mutant W234A had no effects on these properties. Finally, we analyzed the molecular basis of these experimental findings in-silico using the dimer (PDB ID: 1KU0) and the monomer (PDB ID: 2W22) lipase structures. The computational analyses confirmed that W211 stabilized the intermolecular interactions in the dimer lipase and it is critical to the stability of the monomer lipase. Explicitly W211 confers stability to the dimer and the monomer lipase through distinct aromatic interactions with Y273-Y282 and H87-P232 respectively. The insights revealed by this work shed light not only on the mechanism of thermostability and its relation to aggregation but also on the particular role of the conserved lid tryptophan in the thermoalkalophilic lipases. |
| first_indexed | 2024-12-13T20:51:38Z |
| format | Article |
| id | doaj.art-b104159f963c4873bc79fee4b0087c78 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-13T20:51:38Z |
| publishDate | 2013-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-b104159f963c4873bc79fee4b0087c782022-12-21T23:31:51ZengPublic Library of Science (PLoS)PLoS ONE1932-62032013-01-01812e8518610.1371/journal.pone.0085186The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases.Emel TimucinO Ugur SezermanWe hypothesize that aggregation of thermoalkalophilic lipases could be a thermostability mechanism. The conserved tryptophans (W211, W234) in the lid are of particular interest owing to their previous involvements in aggregation and thermostability mechanisms in many other proteins. The thermoalkalophilic lipase from Bacillus thermocatenulatus (BTL2) and its mutants (W211A, W234A) were expressed and purified to homogeneity. We found that, when aggregated, BTL2 is more thermostable than its non-aggregating form, showing that aggregation potentiates thermostability in the thermoalkalophilic lipase. Among the two lid mutants, the W211A lowered aggregation tendency drastically and resulted in a much less thermostable variant of BTL2, which indicated that W211 stabilizes the intermolecular interactions in BTL2 aggregates. Further thermoactivity and CD spectroscopy analyses showed that W211A also led to a strong decrease in the optimal and the melting temperature of BTL2, implying stabilization by W211 also to the intramolecular interactions. The other lid mutant W234A had no effects on these properties. Finally, we analyzed the molecular basis of these experimental findings in-silico using the dimer (PDB ID: 1KU0) and the monomer (PDB ID: 2W22) lipase structures. The computational analyses confirmed that W211 stabilized the intermolecular interactions in the dimer lipase and it is critical to the stability of the monomer lipase. Explicitly W211 confers stability to the dimer and the monomer lipase through distinct aromatic interactions with Y273-Y282 and H87-P232 respectively. The insights revealed by this work shed light not only on the mechanism of thermostability and its relation to aggregation but also on the particular role of the conserved lid tryptophan in the thermoalkalophilic lipases.http://europepmc.org/articles/PMC3877348?pdf=render |
| spellingShingle | Emel Timucin O Ugur Sezerman The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases. PLoS ONE |
| title | The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases. |
| title_full | The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases. |
| title_fullStr | The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases. |
| title_full_unstemmed | The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases. |
| title_short | The conserved lid tryptophan, W211, potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases. |
| title_sort | conserved lid tryptophan w211 potentiates thermostability and thermoactivity in bacterial thermoalkalophilic lipases |
| url | http://europepmc.org/articles/PMC3877348?pdf=render |
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