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The minimal domain of adipose triglyceride lipase (ATGL) ranges until leucine 254 and can be activated and inhibited by CGI-58 and G0S2, respectively.

Cornaciu I, Boeszoermenyi A, Lindermuth H, Nagy HM, Cerk IK, Ebner C, Salzburger B, Gruber A, Schweiger M, Zechner R, Lass A, Zimmermann R, Oberer M - PLoS ONE (2011)

Bottom Line: Yet, neither an experimentally determined 3D structure nor a model of ATGL is currently available, which would help to understand how CGI-58 and G0S2 modulate ATGL's activity.Based on these data, we generated a 3D homology model for the minimal domain.Our data provide insights into the structure-function relationship of ATGL and indicate higher structural similarities in the N-terminal halves of mammalian patatin-like phospholipase domain containing proteins, (PNPLA1, -2,- 3 and -5) than originally anticipated.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biosciences, University of Graz, Graz, Austria.

ABSTRACT
Adipose triglyceride lipase (ATGL) is the rate-limiting enzyme of lipolysis. ATGL specifically hydrolyzes triacylglycerols (TGs), thereby generating diacylglycerols and free fatty acids. ATGL's enzymatic activity is co-activated by the protein comparative gene identification-58 (CGI-58) and inhibited by the protein G0/G1 switch gene 2 (G0S2). The enzyme is predicted to act through a catalytic dyad (Ser47, Asp166) located within the conserved patatin domain (Ile10-Leu178). Yet, neither an experimentally determined 3D structure nor a model of ATGL is currently available, which would help to understand how CGI-58 and G0S2 modulate ATGL's activity. In this study we determined the minimal active domain of ATGL. This minimal fragment of ATGL could still be activated and inhibited by CGI-58 and G0S2, respectively. Furthermore, we show that this minimal domain is sufficient for protein-protein interaction of ATGL with its regulatory proteins. Based on these data, we generated a 3D homology model for the minimal domain. It strengthens our experimental finding that amino acids between Leu178 and Leu254 are essential for the formation of a stable protein domain related to the patatin fold. Our data provide insights into the structure-function relationship of ATGL and indicate higher structural similarities in the N-terminal halves of mammalian patatin-like phospholipase domain containing proteins, (PNPLA1, -2,- 3 and -5) than originally anticipated.

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Model of mouse ATGL254.A. Homology-modeled structure of mouse ATGL ranging from residue Met1-Leu254 in cartoon representation. Residues Trp8-Leu178 are colored in light blue. Ser47 and Asp166, which form the catalytic dyad, are colored by atom; Gly14-Phe17, which are thought to be involved in forming the oxyanion hole are in red. N- and C-termini are indicated. B. Structural alignment of Pat17 (PDB code 1OXW) and cPLA2 (PDB code 1CJY) with the 3D model of ATGL254. Residues at the catalytic site (oxyanion hole, GXSXG motif, catalytic Asp) are highlighted in red. α-helices and β-strands are indicated in green and orange, respectively. Red numbers indicate extra amino acids in the structure of Pat17 and cPLA2. C. Left panel: model of ATGL254; middle panel: overlay of the 3D model of ATGL254 with Pat17 (colored grey and yellow); right panel: structure of Pat17. Val319-Lys383 of Pat17 (yellow) suggest a possible further structural organization of approximately 60 additional residues in mATGL.
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pone-0026349-g007: Model of mouse ATGL254.A. Homology-modeled structure of mouse ATGL ranging from residue Met1-Leu254 in cartoon representation. Residues Trp8-Leu178 are colored in light blue. Ser47 and Asp166, which form the catalytic dyad, are colored by atom; Gly14-Phe17, which are thought to be involved in forming the oxyanion hole are in red. N- and C-termini are indicated. B. Structural alignment of Pat17 (PDB code 1OXW) and cPLA2 (PDB code 1CJY) with the 3D model of ATGL254. Residues at the catalytic site (oxyanion hole, GXSXG motif, catalytic Asp) are highlighted in red. α-helices and β-strands are indicated in green and orange, respectively. Red numbers indicate extra amino acids in the structure of Pat17 and cPLA2. C. Left panel: model of ATGL254; middle panel: overlay of the 3D model of ATGL254 with Pat17 (colored grey and yellow); right panel: structure of Pat17. Val319-Lys383 of Pat17 (yellow) suggest a possible further structural organization of approximately 60 additional residues in mATGL.

Mentions: After establishing the minimal requirements for retaining ATGL's enzymatic activity, we wanted to gain more insight into the structural background enabling the catalytic activity and its protein-protein interactions. Currently, no experimental 3D structure is available for ATGL. Therefore we performed homology modeling of ATGL ranging until residue Leu254. Pat17 and cPLA2, the two patatin family members with known 3D structures, were used as templates. Due to the overall low sequence identity with the templates, the resulting models do not provide atomic details. Therefore it cannot be used for detailed interpretation of the structure, however should be viewed as giving a glimpse of the overall structure of the protein and its domain boundaries. Insights into the potential domain architecture of the N-terminal half of ATGL are given by the model (Figure 7). The template Pat17 is a 386 residue protein. Its crystal structure revealed a single-domain protein, harboring a central β-sheet with two α-helices on the concave side and 7 α-helices on the convex side. Although the patatin fold shares similarities with the canonical α/β hydrolase fold, it also harbors features that are clearly distinct from the α/β hydrolase fold, e.g. a six-stranded sheet as the core, in which five parallel sheets are followed by one anti-parallel strand, and the hydrolytic activity is carried out by a catalytic dyad [14]. In the 3D model of ATGL254, the catalytic dyad residues Ser47 and Asp166 along with a potential oxyanion hole forming residues Gly14-Phe17 are in an overall spatial arrangement which allows enzymatic activity. Residues Asn9-Leu178 in mATGL (blue in Figure 7A and C), which correspond to residues Thr30–Leu227 in Pat17, contribute to 4 of the 6 strands in the central β-sheet. The helical packing on the convex side of the central sheet of the ATGL model allows the formation of a compact arrangement of secondary structure elements. However, residues within this range are only involved in forming one partial α-helix at the concave side of the protein (Figure 7A and C, compare also with Figure 3 in [4]). Therefore it seems very likely, that these residues are not sufficient to build a stable and functional protein domain and that residues beyond Lys179 (cyan in Figure 7A and C) play an integral part in forming the active protein domain. Interestingly, residues Lys180 to Leu254 further complement the central parallel β-sheet and form helices on the concave side of the sheet in our model (colored cyan in Figure 7A). These residues are probably essential for a stable fold of ATGL and play a crucial role in creating the core of this protein domain. In summary, our 3D modeling data are in agreement with our biochemical observations, showing that residues within the N-terminal part of ATGL until Leu254 are essential for forming a stable, active protein domain.


The minimal domain of adipose triglyceride lipase (ATGL) ranges until leucine 254 and can be activated and inhibited by CGI-58 and G0S2, respectively.

Cornaciu I, Boeszoermenyi A, Lindermuth H, Nagy HM, Cerk IK, Ebner C, Salzburger B, Gruber A, Schweiger M, Zechner R, Lass A, Zimmermann R, Oberer M - PLoS ONE (2011)

Model of mouse ATGL254.A. Homology-modeled structure of mouse ATGL ranging from residue Met1-Leu254 in cartoon representation. Residues Trp8-Leu178 are colored in light blue. Ser47 and Asp166, which form the catalytic dyad, are colored by atom; Gly14-Phe17, which are thought to be involved in forming the oxyanion hole are in red. N- and C-termini are indicated. B. Structural alignment of Pat17 (PDB code 1OXW) and cPLA2 (PDB code 1CJY) with the 3D model of ATGL254. Residues at the catalytic site (oxyanion hole, GXSXG motif, catalytic Asp) are highlighted in red. α-helices and β-strands are indicated in green and orange, respectively. Red numbers indicate extra amino acids in the structure of Pat17 and cPLA2. C. Left panel: model of ATGL254; middle panel: overlay of the 3D model of ATGL254 with Pat17 (colored grey and yellow); right panel: structure of Pat17. Val319-Lys383 of Pat17 (yellow) suggest a possible further structural organization of approximately 60 additional residues in mATGL.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198459&req=5

pone-0026349-g007: Model of mouse ATGL254.A. Homology-modeled structure of mouse ATGL ranging from residue Met1-Leu254 in cartoon representation. Residues Trp8-Leu178 are colored in light blue. Ser47 and Asp166, which form the catalytic dyad, are colored by atom; Gly14-Phe17, which are thought to be involved in forming the oxyanion hole are in red. N- and C-termini are indicated. B. Structural alignment of Pat17 (PDB code 1OXW) and cPLA2 (PDB code 1CJY) with the 3D model of ATGL254. Residues at the catalytic site (oxyanion hole, GXSXG motif, catalytic Asp) are highlighted in red. α-helices and β-strands are indicated in green and orange, respectively. Red numbers indicate extra amino acids in the structure of Pat17 and cPLA2. C. Left panel: model of ATGL254; middle panel: overlay of the 3D model of ATGL254 with Pat17 (colored grey and yellow); right panel: structure of Pat17. Val319-Lys383 of Pat17 (yellow) suggest a possible further structural organization of approximately 60 additional residues in mATGL.
Mentions: After establishing the minimal requirements for retaining ATGL's enzymatic activity, we wanted to gain more insight into the structural background enabling the catalytic activity and its protein-protein interactions. Currently, no experimental 3D structure is available for ATGL. Therefore we performed homology modeling of ATGL ranging until residue Leu254. Pat17 and cPLA2, the two patatin family members with known 3D structures, were used as templates. Due to the overall low sequence identity with the templates, the resulting models do not provide atomic details. Therefore it cannot be used for detailed interpretation of the structure, however should be viewed as giving a glimpse of the overall structure of the protein and its domain boundaries. Insights into the potential domain architecture of the N-terminal half of ATGL are given by the model (Figure 7). The template Pat17 is a 386 residue protein. Its crystal structure revealed a single-domain protein, harboring a central β-sheet with two α-helices on the concave side and 7 α-helices on the convex side. Although the patatin fold shares similarities with the canonical α/β hydrolase fold, it also harbors features that are clearly distinct from the α/β hydrolase fold, e.g. a six-stranded sheet as the core, in which five parallel sheets are followed by one anti-parallel strand, and the hydrolytic activity is carried out by a catalytic dyad [14]. In the 3D model of ATGL254, the catalytic dyad residues Ser47 and Asp166 along with a potential oxyanion hole forming residues Gly14-Phe17 are in an overall spatial arrangement which allows enzymatic activity. Residues Asn9-Leu178 in mATGL (blue in Figure 7A and C), which correspond to residues Thr30–Leu227 in Pat17, contribute to 4 of the 6 strands in the central β-sheet. The helical packing on the convex side of the central sheet of the ATGL model allows the formation of a compact arrangement of secondary structure elements. However, residues within this range are only involved in forming one partial α-helix at the concave side of the protein (Figure 7A and C, compare also with Figure 3 in [4]). Therefore it seems very likely, that these residues are not sufficient to build a stable and functional protein domain and that residues beyond Lys179 (cyan in Figure 7A and C) play an integral part in forming the active protein domain. Interestingly, residues Lys180 to Leu254 further complement the central parallel β-sheet and form helices on the concave side of the sheet in our model (colored cyan in Figure 7A). These residues are probably essential for a stable fold of ATGL and play a crucial role in creating the core of this protein domain. In summary, our 3D modeling data are in agreement with our biochemical observations, showing that residues within the N-terminal part of ATGL until Leu254 are essential for forming a stable, active protein domain.

Bottom Line: Yet, neither an experimentally determined 3D structure nor a model of ATGL is currently available, which would help to understand how CGI-58 and G0S2 modulate ATGL's activity.Based on these data, we generated a 3D homology model for the minimal domain.Our data provide insights into the structure-function relationship of ATGL and indicate higher structural similarities in the N-terminal halves of mammalian patatin-like phospholipase domain containing proteins, (PNPLA1, -2,- 3 and -5) than originally anticipated.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biosciences, University of Graz, Graz, Austria.

ABSTRACT
Adipose triglyceride lipase (ATGL) is the rate-limiting enzyme of lipolysis. ATGL specifically hydrolyzes triacylglycerols (TGs), thereby generating diacylglycerols and free fatty acids. ATGL's enzymatic activity is co-activated by the protein comparative gene identification-58 (CGI-58) and inhibited by the protein G0/G1 switch gene 2 (G0S2). The enzyme is predicted to act through a catalytic dyad (Ser47, Asp166) located within the conserved patatin domain (Ile10-Leu178). Yet, neither an experimentally determined 3D structure nor a model of ATGL is currently available, which would help to understand how CGI-58 and G0S2 modulate ATGL's activity. In this study we determined the minimal active domain of ATGL. This minimal fragment of ATGL could still be activated and inhibited by CGI-58 and G0S2, respectively. Furthermore, we show that this minimal domain is sufficient for protein-protein interaction of ATGL with its regulatory proteins. Based on these data, we generated a 3D homology model for the minimal domain. It strengthens our experimental finding that amino acids between Leu178 and Leu254 are essential for the formation of a stable protein domain related to the patatin fold. Our data provide insights into the structure-function relationship of ATGL and indicate higher structural similarities in the N-terminal halves of mammalian patatin-like phospholipase domain containing proteins, (PNPLA1, -2,- 3 and -5) than originally anticipated.

Show MeSH
Related in: MedlinePlus