N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase.
The gene encoding the TPL N-terminal domain (N-TPL), fused with a His6-tag, was cloned and expressed in Pichia pastoris, under the control of the glyceraldehyde-3-phosphate dehydrogenase (GAP) constitutive promoter. The recombinant protein was successfully expressed and secreted with an expression l...
| Main Authors: | , , , |
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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/PMC3745449?pdf=render |
| _version_ | 1818587384317476864 |
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| author | Madiha Bou Ali Aida Karray Youssef Gargouri Yassine Ben Ali |
| author_facet | Madiha Bou Ali Aida Karray Youssef Gargouri Yassine Ben Ali |
| author_sort | Madiha Bou Ali |
| collection | DOAJ |
| description | The gene encoding the TPL N-terminal domain (N-TPL), fused with a His6-tag, was cloned and expressed in Pichia pastoris, under the control of the glyceraldehyde-3-phosphate dehydrogenase (GAP) constitutive promoter. The recombinant protein was successfully expressed and secreted with an expression level of 5 mg/l of culture medium after 2 days of culture. The N-TPL was purified through a one-step Ni-NTA affinity column with a purification factor of approximately 23-fold. The purified N-TPL, with a molecular mass of 35 kDa, had a specific activity of 70 U/mg on tributyrin. Surprisingly, this domain was able to hydrolyse long chain TG with a specific activity of 11 U/mg using olive oil as substrate. This result was confirmed by TLC analysis showing that the N-TPL was able to hydrolyse insoluble substrates as olive oil. N-TPL was unstable at temperatures over 37°C and lost 70% of its activity at acid pH, after 5 min of incubation. The N-TPL exhibited non linear kinetics, indicating its rapid denaturation at the tributyrin-water interface. Colipase increased the N-TPL stability at the lipid-water interface, so the TPL N-terminal domain probably formed functional interactions with colipase despite the absence of the C-terminal domain. |
| first_indexed | 2024-12-16T09:08:00Z |
| format | Article |
| id | doaj.art-704f5ce0a8ad4ff0b87990c574525319 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-16T09:08:00Z |
| publishDate | 2013-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-704f5ce0a8ad4ff0b87990c5745253192022-12-21T22:37:03ZengPublic Library of Science (PLoS)PLoS ONE1932-62032013-01-0188e7160510.1371/journal.pone.0071605N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase.Madiha Bou AliAida KarrayYoussef GargouriYassine Ben AliThe gene encoding the TPL N-terminal domain (N-TPL), fused with a His6-tag, was cloned and expressed in Pichia pastoris, under the control of the glyceraldehyde-3-phosphate dehydrogenase (GAP) constitutive promoter. The recombinant protein was successfully expressed and secreted with an expression level of 5 mg/l of culture medium after 2 days of culture. The N-TPL was purified through a one-step Ni-NTA affinity column with a purification factor of approximately 23-fold. The purified N-TPL, with a molecular mass of 35 kDa, had a specific activity of 70 U/mg on tributyrin. Surprisingly, this domain was able to hydrolyse long chain TG with a specific activity of 11 U/mg using olive oil as substrate. This result was confirmed by TLC analysis showing that the N-TPL was able to hydrolyse insoluble substrates as olive oil. N-TPL was unstable at temperatures over 37°C and lost 70% of its activity at acid pH, after 5 min of incubation. The N-TPL exhibited non linear kinetics, indicating its rapid denaturation at the tributyrin-water interface. Colipase increased the N-TPL stability at the lipid-water interface, so the TPL N-terminal domain probably formed functional interactions with colipase despite the absence of the C-terminal domain.http://europepmc.org/articles/PMC3745449?pdf=render |
| spellingShingle | Madiha Bou Ali Aida Karray Youssef Gargouri Yassine Ben Ali N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase. PLoS ONE |
| title | N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase. |
| title_full | N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase. |
| title_fullStr | N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase. |
| title_full_unstemmed | N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase. |
| title_short | N-terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase. |
| title_sort | n terminal domain of turkey pancreatic lipase is active on long chain triacylglycerols and stabilized by colipase |
| url | http://europepmc.org/articles/PMC3745449?pdf=render |
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