Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase
Human triacylglycerol hydrolase (hTGH) has been shown to play a role in hepatic lipid metabolism. Triacylglycerol hydrolase (TGH) hydrolyzes insoluble carboxylic esters at lipid/water interfaces, although the mechanism by which the enzyme adsorbs to lipid droplets is unclear. Three-dimensional model...
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| Format: | Article |
| Language: | English |
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Elsevier
2006-02-01
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| Series: | Journal of Lipid Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022227520336385 |
| _version_ | 1818385186010693632 |
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| author | Mustafa Alam Dean Gilham Dennis E. Vance Richard Lehner |
| author_facet | Mustafa Alam Dean Gilham Dennis E. Vance Richard Lehner |
| author_sort | Mustafa Alam |
| collection | DOAJ |
| description | Human triacylglycerol hydrolase (hTGH) has been shown to play a role in hepatic lipid metabolism. Triacylglycerol hydrolase (TGH) hydrolyzes insoluble carboxylic esters at lipid/water interfaces, although the mechanism by which the enzyme adsorbs to lipid droplets is unclear. Three-dimensional modeling of hTGH predicts that catalytic residues are adjacent to an α-helix that may mediate TGH/lipid interaction. The helix contains a putative neutral lipid binding domain consisting of the octapeptide FLDLIADV (amino acid residues 417–424) with the consensus sequence FLXLXXXn (where n is a nonpolar residue and X is any amino acid except proline) identified in several other proteins that bind or metabolize neutral lipids. Deletion of this α-helix abolished the lipolytic activity of hTGH. Replacement of F417 with alanine reduced activity by 40% toward both insoluble and soluble esters, whereas replacement of L418 and L420 with alanine did not. Another potential mechanism of increasing TGH affinity for lipid is via reversible acylation. Molecular modeling predicts that C390 is available for covalent acylation. However, neither chemical modification of C390 nor mutation to alanine affected activity. Our findings indicate that F417 but not L418, L420, or C390 participates in substrate hydrolysis by hTGH. |
| first_indexed | 2024-12-14T03:34:09Z |
| format | Article |
| id | doaj.art-a0d455e3896b4ce49a58d2d887062c1f |
| institution | Directory Open Access Journal |
| issn | 0022-2275 |
| language | English |
| last_indexed | 2024-12-14T03:34:09Z |
| publishDate | 2006-02-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Lipid Research |
| spelling | doaj.art-a0d455e3896b4ce49a58d2d887062c1f2022-12-21T23:18:40ZengElsevierJournal of Lipid Research0022-22752006-02-01472375383Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolaseMustafa Alam0Dean Gilham1Dennis E. Vance2Richard Lehner3Department of Pediatrics, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Cell Biology, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Biochemistry, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2Department of Pediatrics, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Cell Biology, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Biochemistry, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2Department of Pediatrics, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Cell Biology, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Biochemistry, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2Department of Pediatrics, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Cell Biology, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2; Department of Biochemistry, Canadian Institutes of Health Research Group on the Molecular and Cell Biology of Lipids, University of Alberta, Edmonton, Alberta, Canada T6G 2S2Human triacylglycerol hydrolase (hTGH) has been shown to play a role in hepatic lipid metabolism. Triacylglycerol hydrolase (TGH) hydrolyzes insoluble carboxylic esters at lipid/water interfaces, although the mechanism by which the enzyme adsorbs to lipid droplets is unclear. Three-dimensional modeling of hTGH predicts that catalytic residues are adjacent to an α-helix that may mediate TGH/lipid interaction. The helix contains a putative neutral lipid binding domain consisting of the octapeptide FLDLIADV (amino acid residues 417–424) with the consensus sequence FLXLXXXn (where n is a nonpolar residue and X is any amino acid except proline) identified in several other proteins that bind or metabolize neutral lipids. Deletion of this α-helix abolished the lipolytic activity of hTGH. Replacement of F417 with alanine reduced activity by 40% toward both insoluble and soluble esters, whereas replacement of L418 and L420 with alanine did not. Another potential mechanism of increasing TGH affinity for lipid is via reversible acylation. Molecular modeling predicts that C390 is available for covalent acylation. However, neither chemical modification of C390 nor mutation to alanine affected activity. Our findings indicate that F417 but not L418, L420, or C390 participates in substrate hydrolysis by hTGH.http://www.sciencedirect.com/science/article/pii/S0022227520336385lipolysismutagenesiscarboxylesteraselipase |
| spellingShingle | Mustafa Alam Dean Gilham Dennis E. Vance Richard Lehner Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase Journal of Lipid Research lipolysis mutagenesis carboxylesterase lipase |
| title | Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase |
| title_full | Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase |
| title_fullStr | Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase |
| title_full_unstemmed | Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase |
| title_short | Mutation of F417 but not of L418 or L420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase |
| title_sort | mutation of f417 but not of l418 or l420 in the lipid binding domain decreases the activity of triacylglycerol hydrolase |
| topic | lipolysis mutagenesis carboxylesterase lipase |
| url | http://www.sciencedirect.com/science/article/pii/S0022227520336385 |
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