The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene
The alkaline cold-active lipase from Pseudomonas fluorescens AMS8 undergoes major structural changes when reacted with hydrophobic organic solvents. In toluene, the AMS8 lipase catalytic region is exposed by the moving hydrophobic lid 2 (Glu-148 to Gly-167). Solvent-accessible surface area analysis...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI
2017
|
| Online Access: | http://psasir.upm.edu.my/id/eprint/63526/1/The%20Role%20of%20Solvent-Accessible%20Leu-208%20of%20Cold-Active%20Pseudomonas%20fluorescens%20Strain%20AMS8%20Lipase%20in%20Interfacial%20Activation.pdf |
| _version_ | 1796977758576836608 |
|---|---|
| author | Yaacob, Norhayati Kamarudin, Nor Hafizah Ahmad Thean, Adam Chor Leow Salleh, Abu Bakar Rahman, Raja Noor Zaliha Raja Abd Ali, Mohd. Shukuri Mohamad |
| author_facet | Yaacob, Norhayati Kamarudin, Nor Hafizah Ahmad Thean, Adam Chor Leow Salleh, Abu Bakar Rahman, Raja Noor Zaliha Raja Abd Ali, Mohd. Shukuri Mohamad |
| author_sort | Yaacob, Norhayati |
| collection | UPM |
| description | The alkaline cold-active lipase from Pseudomonas fluorescens AMS8 undergoes major structural changes when reacted with hydrophobic organic solvents. In toluene, the AMS8 lipase catalytic region is exposed by the moving hydrophobic lid 2 (Glu-148 to Gly-167). Solvent-accessible surface area analysis revealed that Leu-208, which is located next to the nucleophilic Ser-207 has a focal function in influencing substrate accessibility and flexibility of the catalytic pocket. Based on molecular dynamic simulations, it was found that Leu-208 strongly facilitates the lid 2 opening via its side-chain. The KM and Kcat/KM of L208A mutant were substrate dependent as it preferred a smaller-chain ester (pNP-caprylate) as compared to medium (pNP-laurate) or long-chain (pNP-palmitate) esters. In esterification of ethyl hexanoate, L208A promotes a higher ester conversion rate at 20 °C but not at 30 °C, as a 27% decline was observed. Interestingly, the wild-type (WT) lipase's conversion rate was found to increase with a higher temperature. WT lipase AMS8 esterification was higher in toluene as compared to L208A. Hence, the results showed that Leu-208 of AMS8 lipase plays an important role in steering a broad range of substrates into its active site region by regulating the flexibility of this region. Leu-208 is therefore predicted to be crucial for its role in interfacial activation and catalysis in toluene. |
| first_indexed | 2024-03-06T09:44:46Z |
| format | Article |
| id | upm.eprints-63526 |
| institution | Universiti Putra Malaysia |
| language | English |
| last_indexed | 2024-03-06T09:44:46Z |
| publishDate | 2017 |
| publisher | MDPI |
| record_format | dspace |
| spelling | upm.eprints-635262018-11-05T01:10:01Z http://psasir.upm.edu.my/id/eprint/63526/ The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene Yaacob, Norhayati Kamarudin, Nor Hafizah Ahmad Thean, Adam Chor Leow Salleh, Abu Bakar Rahman, Raja Noor Zaliha Raja Abd Ali, Mohd. Shukuri Mohamad The alkaline cold-active lipase from Pseudomonas fluorescens AMS8 undergoes major structural changes when reacted with hydrophobic organic solvents. In toluene, the AMS8 lipase catalytic region is exposed by the moving hydrophobic lid 2 (Glu-148 to Gly-167). Solvent-accessible surface area analysis revealed that Leu-208, which is located next to the nucleophilic Ser-207 has a focal function in influencing substrate accessibility and flexibility of the catalytic pocket. Based on molecular dynamic simulations, it was found that Leu-208 strongly facilitates the lid 2 opening via its side-chain. The KM and Kcat/KM of L208A mutant were substrate dependent as it preferred a smaller-chain ester (pNP-caprylate) as compared to medium (pNP-laurate) or long-chain (pNP-palmitate) esters. In esterification of ethyl hexanoate, L208A promotes a higher ester conversion rate at 20 °C but not at 30 °C, as a 27% decline was observed. Interestingly, the wild-type (WT) lipase's conversion rate was found to increase with a higher temperature. WT lipase AMS8 esterification was higher in toluene as compared to L208A. Hence, the results showed that Leu-208 of AMS8 lipase plays an important role in steering a broad range of substrates into its active site region by regulating the flexibility of this region. Leu-208 is therefore predicted to be crucial for its role in interfacial activation and catalysis in toluene. MDPI 2017-08 Article PeerReviewed text en http://psasir.upm.edu.my/id/eprint/63526/1/The%20Role%20of%20Solvent-Accessible%20Leu-208%20of%20Cold-Active%20Pseudomonas%20fluorescens%20Strain%20AMS8%20Lipase%20in%20Interfacial%20Activation.pdf Yaacob, Norhayati and Kamarudin, Nor Hafizah Ahmad and Thean, Adam Chor Leow and Salleh, Abu Bakar and Rahman, Raja Noor Zaliha Raja Abd and Ali, Mohd. Shukuri Mohamad (2017) The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene. Molecules, 22 (8). pp. 1-20. ISSN 1420-3049 https://www.mdpi.com/1420-3049/22/8/1312/xml 10.3390/molecules22081312 |
| spellingShingle | Yaacob, Norhayati Kamarudin, Nor Hafizah Ahmad Thean, Adam Chor Leow Salleh, Abu Bakar Rahman, Raja Noor Zaliha Raja Abd Ali, Mohd. Shukuri Mohamad The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene |
| title | The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene |
| title_full | The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene |
| title_fullStr | The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene |
| title_full_unstemmed | The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene |
| title_short | The role of solvent-accessible leu-208 of cold-active Pseudomonas fluorescens strain AMS8 lipase in interfacial activation, substrate accessibility and low-molecular weight esterification in the presence of toluene |
| title_sort | role of solvent accessible leu 208 of cold active pseudomonas fluorescens strain ams8 lipase in interfacial activation substrate accessibility and low molecular weight esterification in the presence of toluene |
| url | http://psasir.upm.edu.my/id/eprint/63526/1/The%20Role%20of%20Solvent-Accessible%20Leu-208%20of%20Cold-Active%20Pseudomonas%20fluorescens%20Strain%20AMS8%20Lipase%20in%20Interfacial%20Activation.pdf |
| work_keys_str_mv | AT yaacobnorhayati theroleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT kamarudinnorhafizahahmad theroleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT theanadamchorleow theroleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT sallehabubakar theroleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT rahmanrajanoorzaliharajaabd theroleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT alimohdshukurimohamad theroleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT yaacobnorhayati roleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT kamarudinnorhafizahahmad roleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT theanadamchorleow roleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT sallehabubakar roleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT rahmanrajanoorzaliharajaabd roleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene AT alimohdshukurimohamad roleofsolventaccessibleleu208ofcoldactivepseudomonasfluorescensstrainams8lipaseininterfacialactivationsubstrateaccessibilityandlowmolecularweightesterificationinthepresenceoftoluene |