This entry represents a family of eukaryotic lipases, including gastric triacylglycerol lipase (LIPF) lysosomal acid lipase (LIPA) and LIPJ, LIPK, LIPM, LIPN. Mutations in LIPA cause Wolman disease, Cholesterol Ester Storage Disease, CESD and mutation in LIPN causes Late-Onset Form of Autosomal-Recessive Congenital Ichthyosis
17 moreTitle: Comparative studies of mammalian acid lipases: Evidence for a new gene family in mouse and rat (Lipo) Holmes RS, Cox LA, Vandeberg JL Ref: Comparative Biochemistry & Physiology Part D Genomics Proteomics, 5:217, 2010 : PubMed
At least six families of mammalian acid lipases (E.C. 3.1.1.-) catalyse the hydrolysis of triglycerides in the body, designated as LIPA (lysosomal), LIPF (gastric), LIPJ (testis) and LIPK, LIPM and LIPN (epidermal), which belong to the AB hydrolase superfamily. In this study, in silico methods were used to predict the amino acid sequences, secondary and tertiary structures, and gene locations for acid lipase genes and encoded proteins using data from several mammalian genome projects. Mammalian acid lipase genes were located within a gene cluster for each of the 8 mammalian genomes examined, including human (Homo sapiens), chimpanzee (Pons troglodytes), rhesus monkey (Macacca mulatta), mouse (Mus musculus), rat (Rattus norvegicus), cow (Bos taurus), horse (Equus caballus) and dog (Canis familaris), with each containing 9 coding exons. Human and mouse acid lipases shared 44-87% sequence identity and exhibited sequence alignments and identities for key amino acid residues and conservation of predicted secondary and tertiary structures with those previously reported for human gastric lipase (LIPF) (Roussel et al., 1999). Evidence for a new family of acid lipase genes is reported for mouse and rat genomes, designated as Lipo. Mouse acid lipase genes are subject to differential mRNA tissue expression, with Lipa showing wide tissue expression, while others have a more restricted tissue expression in the digestive tract (Lipf), salivary gland (Lipo) and epidermal tissues (Lipk, Lipm and Lipn). Phylogenetic analyses of the mammalian acid lipase gene families suggested that these genes are products of gene duplication events prior to eutherian mammalian evolution and derived from an ancestral vertebrate LIPA gene, which is present in the frog, Xenopus tropicalis.
Fat digestion in humans requires not only the classical pancreatic lipase but also gastric lipase, which is stable and active despite the highly acidic stomach environment. We have solved the structure of recombinant human gastric lipase at 3.0 A resolution, the first structure to be described within the mammalian acid lipase family. This globular enzyme (379 residues) consists of a core domain, belonging to the alpha/beta hydrolase fold family, and an extrusion domain. It possesses a classical catalytic triad (Ser 153, His 353, Asp 324) and an oxyanion hole (NH groups of Gln 154 and Leu 67). Four N-glycosylation sites were identified on the electron density maps. The catalytic serine is deeply buried under the extrusion domain, which is composed of a 'cap' domain and a segment consisting of 30 residues, which can be defined as a lid. Its displacement is necessary for the substrates to access the active site. A phosphonate inhibitor was positioned in the active site which clearly suggests the location of the hydrophobic substrate binding site.
Fat digestion in humans requires not only the classical pancreatic lipase but also gastric lipase, which is stable and active despite the highly acidic stomach environment. We report here the structure of recombinant human gastric lipase at 3.0-A resolution, the first structure to be described within the mammalian acid lipase family. This globular enzyme (379 residues) consists of a core domain belonging to the alpha/beta hydrolase-fold family and a "cap" domain, which is analogous to that present in serine carboxypeptidases. It possesses a classical catalytic triad (Ser-153, His-353, Asp-324) and an oxyanion hole (NH groups of Gln-154 and Leu-67). Four N-glycosylation sites were identified on the electron density maps. The catalytic serine is deeply buried under a segment consisting of 30 residues, which can be defined as a lid and belonging to the cap domain. The displacement of the lid is necessary for the substrates to have access to Ser-153. A phosphonate inhibitor was positioned in the active site that clearly suggests the location of the hydrophobic substrate binding site. The lysosomal acid lipase was modeled by homology, and possible explanations for some previously reported mutations leading to the cholesterol ester storage disease are given based on the present model.
Lysosomal acid lipase (LAL) is the only known enzyme, which hydrolyzes cholesteryl esters and triacylglycerols in lysosomes of multiple cells and tissues. Here, we explored the role of LAL in brown adipose tissue (BAT). LAL-deficient (Lal-/-) mice exhibit markedly reduced UCP1 expression in BAT, modified BAT morphology with accumulation of lysosomes, and mitochondrial dysfunction, consequently leading to regular hypothermic events in mice kept at room temperature. Cold exposure resulted in reduced lipid uptake into BAT, thereby aggravating dyslipidemia and causing life threatening hypothermia in Lal-/- mice. Linking LAL as a potential regulator of lipoprotein lipase activity, we found Angptl4 mRNA expression upregulated in BAT. Our data demonstrate that LAL is critical for shuttling fatty acids derived from circulating lipoproteins to BAT during cold exposure. We conclude that inhibited lysosomal lipid hydrolysis in BAT leads to impaired thermogenesis in Lal-/- mice.
Degradation of lysosomal lipids requires lysosomal acid lipase (LAL), the only intracellular lipase known to be active at acidic pH. We found LAL to be expressed in murine immune cells with highest mRNA expression in macrophages and neutrophils. Furthermore, we observed that loss of LAL in mice caused lipid accumulation in white blood cells in the peripheral circulation, which increased in response to an acute inflammatory stimulus. Lal-deficient (-/-) macrophages accumulate neutral lipids, mainly cholesteryl esters, within lysosomes. The cholesteryl ester fraction is particularly enriched in the PUFAs 18:2 and 20:4, important precursor molecules for lipid mediator synthesis. To investigate whether loss of LAL activity affects the generation of lipid mediators and to eliminate potential systemic effects from other cells and tissues involved in the pronounced phenotype of Lal-/- mice, we treated macrophages from Wt mice with the LAL-specific inhibitor LAListat-2. Acute inhibition of LAL resulted in reduced release of 18:2- and 20:4-derived mediators from macrophages, indicating that lipid hydrolysis by LAL is an important source for lipid mediator synthesis in macrophages. We conclude that lysosomes should be considered as organelles that provide precursor molecules for lipid mediators such as eicosanoids.
        
Title: Endothelial Rab7 GTPase mediates tumor growth and metastasis in lysosomal acid lipase-deficient mice Zhao T, Ding X, Yan C, Du H Ref: Journal of Biological Chemistry, 292:19198, 2017 : PubMed
Tumors depend on their microenvironment for sustained growth, invasion, and metastasis. In this environment, endothelial cells (ECs) are an important stromal cell type interacting with malignant cells to facilitate tumor angiogenesis and cancer cell extravasation. Of note, lysosomal acid lipase (LAL) deficiency facilitates melanoma growth and metastasis. ECs from LAL-deficient (lal(-/-)) mice possess enhanced proliferation, migration, and permeability of inflammatory cells by activating the mammalian target of rapamycin (mTOR) pathway. Here we report that lal(-/-) ECs facilitated in vivo tumor angiogenesis, growth, and metastasis, largely by stimulating tumor cell proliferation, migration, adhesion, and transendothelial migration via increased expression of IL-6 and monocyte chemoattractant protein 1 (MCP-1). This prompted us to look for lysosomal proteins that are involved in lal(-/-) EC dysfunctions. We found that lal(-/-) ECs displayed increased expression of Rab7, a late endosome/lysosome-associated small GTPase. Moreover, Rab7 and mTOR were co-increased and co-localized to lysosomes and physically interacted in lal(-/-) ECs. Rab7 inhibition reversed lal(-/-) EC dysfunctions, including decreasing their enhanced migration and permeability of tumor-stimulatory myeloid cells, and suppressed EC-mediated stimulation of in vitro tumor cell transmigration, proliferation, and migration and in vivo tumor growth and metastasis. Finally, Rab7 inhibition reduced overproduction of reactive oxygen species and increased IL-6 and MCP-1 secretion in lal(-/-) ECs. Our results indicate that metabolic reprogramming resulting from LAL deficiency enhances the ability of ECs to stimulate tumor cell proliferation and metastasis through stimulation of lysosome-anchored Rab7 activity.
Lysosomal acid lipase (LAL) is essential for the clearance of endocytosed cholesteryl ester and triglyceride-rich chylomicron remnants. Humans and mice with defective or absent LAL activity accumulate large amounts of cholesteryl esters and triglycerides in multiple tissues. Although chylomicrons also contain retinyl esters (REs), a role of LAL in the clearance of endocytosed REs has not been reported. In this study, we found that murine LAL exhibits RE hydrolase activity. Pharmacological inhibition of LAL in the human hepatocyte cell line HepG2, incubated with chylomicrons, led to increased accumulation of REs in endosomal/lysosomal fractions. Furthermore, pharmacological inhibition or genetic ablation of LAL in murine liver largely reduced in vitro acid RE hydrolase activity. Interestingly, LAL-deficient mice exhibited increased RE content in the duodenum and jejunum but decreased RE content in the liver. Furthermore, LAL-deficient mice challenged with RE gavage exhibited largely reduced post-prandial circulating RE content, indicating that LAL is required for efficient nutritional vitamin A availability. In summary, our results indicate that LAL is the major acid RE hydrolase and required for functional retinoid homeostasis.
        
Title: Hepatocyte-Specific Expression of Human Lysosome Acid Lipase Corrects Liver Inflammation and Tumor Metastasis in lal(-/-) Mice Du H, Zhao T, Ding X, Yan C Ref: American Journal of Pathology, 185:2379, 2015 : PubMed
The liver is a major organ for lipid synthesis and metabolism. Deficiency of lysosomal acid lipase (LAL; official name Lipa, encoded by Lipa) in mice (lal(-/-)) results in enlarged liver size due to neutral lipid storage in hepatocytes and Kupffer cells. To test the functional role of LAL in hepatocyte, hepatocyte-specific expression of human LAL (hLAL) in lal(-/-) mice was established by cross-breeding of liver-activated promoter (LAP)-driven tTA transgene and (tetO)7-CMV-hLAL transgene with lal(-/-) knockout (KO) (LAP-Tg/KO) triple mice. Hepatocyte-specific expression of hLAL in LAP-Tg/KO triple mice reduced the liver size to the normal level by decreasing lipid storage in both hepatocytes and Kupffer cells. hLAL expression reduced tumor-promoting myeloid-derived suppressive cells in the liver of lal(-/-) mice. As a result, B16 melanoma metastasis to the liver was almost completely blocked. Expression and secretion of multiple tumor-promoting cytokines or chemokines in the liver were also significantly reduced. Because hLAL is a secretory protein, lal(-/-) phenotypes in other compartments (eg, blood, spleen, and lung) also ameliorated, including systemic reduction of myeloid-derived suppressive cells, an increase in CD4(+) and CD8(+) T and B lymphocytes, and reduced B16 melanoma metastasis in the lung. These results support a concept that LAL in hepatocytes is a critical metabolic enzyme in controlling neutral lipid metabolism, liver homeostasis, immune response, and tumor metastasis.
Lysosomal acid lipase (LAL) is an essential enzyme that hydrolyzes triglycerides (TG) and cholesteryl esters (CE) in lysosomes. Mutations of the LIPA gene lead to Wolman disease (WD) and cholesterol ester storage disease (CESD). The disease hallmarks include hepatosplenomegaly and extensive storage of CE and/or TG. The effects of intravenous investigational enzyme therapy (ET) on survival and efficacy were evaluated in Lipa knock out, lal-/- mice with advanced disease using recombinant human LAL (rhLAL). Comparative ET was conducted with lower doses (weekly, 0.8 and 3.2mg/kg) beginning at 16 weeks (study 1), and with higher dose (10mg/kg) in early (8-weeks), middle (16-weeks) and late (24-weeks) disease stages (study 2). In study 1, rhLAL extended the life span of lal-/- mice in a dose dependent manner by 52 (0.8 mg/kg) or 94 (3.2mg/kg) days. This was accompanied by partial correction of cholesterol and TG levels in spleen and liver. In study 2, the high dose resulted in a significant improvement in organ size (liver, spleen and small intestine) and tissue histology as well as significant decreases in cholesterol and TG in all three groups. In the treated livers and spleens the cholesterol and TG levels were reduced to below treatment initiation levels indicating a reversal of disease manifestations, even in advanced disease. ET diminished liver fibrosis and macrophage proliferation. These results show that LAL deficiency can be improved biochemically and histopathologically by various dosages of ET, even in advanced disease.
Autosomal-recessive congenital ichthyoses represent a large and heterogeneous group of disorders of epidermal cornification. Recent data suggest that most of these disorders might result from defective lipid transport and metabolism. In the present study, we describe a late-onset form of recessive ichthyosis in a large consanguineous pedigree. By using a combination of homozygosity mapping and positional candidate-gene screening, we identified a 2 bp deletion in LIPN that segregated with the disease phenotype throughout the family. LIPN encodes one of six acid lipases known to be involved in triglyceride metabolism in mammals . LIPN was found to be exclusively expressed in the epidermis and to be strongly induced during keratinocyte differentiation.
Lipoprotein cholesterol taken up by cells is processed in the endosomal/lysosomal (E/L) compartment by the sequential action of lysosomal acid lipase (LAL), Niemann-Pick C2 (NPC2), and Niemann-Pick C1 (NPC1). Inactivation of NPC2 in mouse caused sequestration of unesterified cholesterol (UC) and expanded the whole animal sterol pool from 2,305 to 4,337 mg/kg. However, this pool increased to 5,408 and 9,480 mg/kg, respectively, when NPC1 or LAL function was absent. The transport defect in mutants lacking NPC2 or NPC1, but not in those lacking LAL, was reversed by cyclodextrin (CD), and the ED(5)(0) values for this reversal varied from ~40 mg/kg in kidney to >20,000 mg/kg in brain in both groups. This reversal occurred only with a CD that could interact with UC. Further, a CD that could interact with, but not solubilize, UC still overcame the transport defect. These studies showed that processing and export of sterol from the late E/L compartment was quantitatively different in mice lacking LAL, NPC2, or NPC1 function. In both npc2(-/-) and npc1(-/-) mice, the transport defect was reversed by a CD that interacted with UC, likely at the membrane/bulk-water interface, allowing sterol to move rapidly to the export site of the E/L compartment.
        
Title: Comparative studies of mammalian acid lipases: Evidence for a new gene family in mouse and rat (Lipo) Holmes RS, Cox LA, Vandeberg JL Ref: Comparative Biochemistry & Physiology Part D Genomics Proteomics, 5:217, 2010 : PubMed
At least six families of mammalian acid lipases (E.C. 3.1.1.-) catalyse the hydrolysis of triglycerides in the body, designated as LIPA (lysosomal), LIPF (gastric), LIPJ (testis) and LIPK, LIPM and LIPN (epidermal), which belong to the AB hydrolase superfamily. In this study, in silico methods were used to predict the amino acid sequences, secondary and tertiary structures, and gene locations for acid lipase genes and encoded proteins using data from several mammalian genome projects. Mammalian acid lipase genes were located within a gene cluster for each of the 8 mammalian genomes examined, including human (Homo sapiens), chimpanzee (Pons troglodytes), rhesus monkey (Macacca mulatta), mouse (Mus musculus), rat (Rattus norvegicus), cow (Bos taurus), horse (Equus caballus) and dog (Canis familaris), with each containing 9 coding exons. Human and mouse acid lipases shared 44-87% sequence identity and exhibited sequence alignments and identities for key amino acid residues and conservation of predicted secondary and tertiary structures with those previously reported for human gastric lipase (LIPF) (Roussel et al., 1999). Evidence for a new family of acid lipase genes is reported for mouse and rat genomes, designated as Lipo. Mouse acid lipase genes are subject to differential mRNA tissue expression, with Lipa showing wide tissue expression, while others have a more restricted tissue expression in the digestive tract (Lipf), salivary gland (Lipo) and epidermal tissues (Lipk, Lipm and Lipn). Phylogenetic analyses of the mammalian acid lipase gene families suggested that these genes are products of gene duplication events prior to eutherian mammalian evolution and derived from an ancestral vertebrate LIPA gene, which is present in the frog, Xenopus tropicalis.
        
Title: Critical roles of lysosomal acid lipase in myelopoiesis Qu P, Shelley WC, Yoder MC, Wu L, Du H, Yan C Ref: American Journal of Pathology, 176:2394, 2010 : PubMed
Lysosomal acid lipase (LAL) is a key enzyme that cleaves cholesteryl esters and triglycerides to generate free fatty acids and cholesterol in lysosomes. Genetic ablation of the lal gene (lal(-/-)) in mice has resulted in a systemic increase of macrophages and neutrophils, causing severe inflammation and pathogenesis in multiple organs. We hypothesized that aberrant growth and differentiation of myeloid cells in lal(-/-) mice arises from dysregulated production of progenitor cells in the bone marrow. Indeed, lal(-/-) mice displayed increased numbers of primitive lin(-)Sca-1(+)c-Kit(+) (LSK) cells and granulocyte-macrophage precursors (GMP). Increased high proliferative potential colony-forming cells (HPP-CFC) were enumerated from cultured lal(-/-) bone marrow cells, as were significantly more CFU-GM, CFU-G, and CFU-M colonies. As a consequence, lal(-/-) mice developed significant myeloid infiltration, particularly with CD11b+/Gr-1+ myeloid-derived suppressive cells in multiple organs. Both decreased apoptosis and increased proliferation contribute to the systemic increase of myeloid cells in lal(-/-) myeloid cells. These lal(-/-) CD11b(+)/Gr-1(+) cells displayed suppressive activity on T cell proliferation and function in vitro. Bone marrow chimeras confirmed that the myeloproliferative disorder in lal(-/-) mice was primarily attributable to autonomous defects in myeloid progenitor cells, although the hematopoietic microenvironment in the lal(-/-) mice did not support hematopoiesis normally. These results provide evidence that LAL is an important regulator of myelopoiesis during hematopoietic development, differentiation, and homeostasis.
        
Title: Critical roles of lysosomal acid lipase in T cell development and function Qu P, Du H, Wilkes DS, Yan C Ref: American Journal of Pathology, 174:944, 2009 : PubMed
Lysosomal acid lipase (LAL) cleaves cholesteryl esters and triglycerides to generate free fatty acids and cholesterol in lysosomes. In LAL gene-knockout (lal(-/-)) mice, blockage of cholesteryl ester and triglyceride metabolism led to abnormal organization of the thymus and spleen, as well as neutral lipid accumulation in these organs. LAL deficiency impaired T cell development in the thymus. Peripheral T cells were reduced dramatically in lal(-/-) mice, due largely to increased apoptosis and decreased proliferation of lal(-/-) T cells in the thymus and peripheral compartments. These lal(-/-) T cells lost the ability to respond to T cell receptor stimulation, including reduced expression of cell surface receptor CD69, abolishment of T cell proliferation, and decreased expression of T lymphokines after stimulation by either anti-CD3 plus anti-CD28 or phorbol-12-myristate-13-acetate and ionomycin. Differentiation of Th1 and Th2 CD4(+) effector lymphocytes by T cell receptor stimulation was blocked in lal(-/-) mice. The ratio of CD4(+)CD25(+)FoxP3(+) Tregs to CD4(+) T cells was increased in lal(-/-) spleens. Bone marrow chimeras demonstrated retardation of T cell development and maturation in lal(-/-) mice due to defects in T cell precursors. Therefore, LAL, its downstream genes, and lipid mediators all play essential roles in development, homeostasis, and function of T cells. The altered development and function of lal(-/-) T cells contributes to disease formation in various organs during LAL deficiency.
        
Title: Lysosomal acid lipase over-expression disrupts lamellar body genesis and alveolar structure in the lung Li Y, Qin Y, Li H, Wu R, Yan C, Du H Ref: International Journal of Experimental Pathology, 88:427, 2007 : PubMed
The functional role of neutral lipids in the lung is poorly understood. Lysosomal acid lipase (LAL) is a critical enzyme in hydrolysis of cholesteryl esters and triglycerides to generate free fatty acids and cholesterol in lysosomes. Human LAL was over-expressed in a doxycycline-controlled system in mouse respiratory epithelial cells to accelerate intracellular neutral lipid degradation and perturb the surfactant homeostasis in the lung. In this animal system, neutral lipid concentrations of pulmonary surfactant were reduced in bronchoalveolar lavage fluid (BALF) in association with decrease of surfactant protein C (SP-C) gene expression. The size and the number of lamellar bodies in alveolar type II epithelial cells (AT II cells) were significantly reduced accordingly. The number of macrophages required for surfactant recycling in BALF was also significantly reduced. As a result of these combinatory effects, emphysema of the alveolar structure was observed. Taken together, neutral lipid homeostasis is essential for maintenance of lamellar body genesis and the alveolar structure in the lung.
Triacylglycerol (TAG) lipases have been thoroughly characterized in mammals and microorganisms. By contrast, very little is known on plant TAG lipases. An Arabidopsis cDNA called AtLip1 (At2g15230), which exhibits strong homology to lysosomal acid lipase, was found to drive the synthesis of an active TAG lipase when expressed in the baculovirus system. The lipase had a maximal activity at pH 6 and the specific activity was estimated to be about 45 micromol min(-1) mg(-1) protein using triolein as a substrate. Knock-out mutant analysis showed no phenotype during germination indicating that this enzyme is fully dispensable for TAG storage breakdown during germination. Northern blot analyses indicated that the transcript is present in all tissues tested.
        
Title: Neutral lipids and peroxisome proliferator-activated receptor-{gamma} control pulmonary gene expression and inflammation-triggered pathogenesis in lysosomal acid lipase knockout mice Lian X, Yan C, Qin Y, Knox L, Li T, Du H Ref: American Journal of Pathology, 167:813, 2005 : PubMed
The functional roles of neutral lipids in the lung are poorly understood. However, blocking cholesteryl ester and triglyceride metabolism in lysosomal acid lipase gene knockout mice (lal-/-) results in severe pathogenic phenotypes in the lung, including massive neutrophil infiltration, foamy macrophage accumulation, unwanted cell growth, and emphysema. To elucidate the mechanism underlining these pathologies, we performed Affymetrix GeneChip microarray analysis of 1-, 3-, and 6-month-old mice and identified aberrant gene expression that progressed with age. Among changed genes, matrix metalloproteinase (MMP)-12, apoptosis inhibitor 6 (Api-6), erythroblast transformation-specific domain (Ets) transcription factor family member Spi-C, and oncogene MafB were increased 100-, 70-, 40-, and 10-fold, respectively, in lal-/- lungs versus the wild-type lungs. The pathogenic increases of these molecules occurred primarily in alveolar type II epithelial cells. Transcriptional activities of the MMP-12 and Api-6 promoters were stimulated by Spi-C or MafB in respiratory epithelial cells. Treatment with 9-hydroxyoctadecanoic acids and ciglitazone significantly rescued lal-/- pulmonary inflammation and aberrant gene expression. In addition, both compounds as well as peroxisome proliferator-activated receptor gamma inhibited MMP-12 and Api-6 promoter activities. These data suggest that inflammation-triggered cell growth and emphysema during lysosomal acid lipase deficiency are partially caused by peroxisome proliferator-activated receptor-gamma inactivation.
        
Title: Lysosomal acid lipase deficiency causes respiratory inflammation and destruction in the lung Lian X, Yan C, Yang L, Xu Y, Du H Ref: American Journal of Physiology Lung Cell Mol Physiol, 286:L801, 2004 : PubMed
The functional roles of neutral lipids are poorly understood in the lung. Blocking cholesteryl ester and triglyceride metabolism in lysosomal acid lipase gene knockout mice (lal-/-) resulted in a high level of neutrophil influx in the lungs as early as 2 mo of age. Bronchoalveolar macrophages appeared foamy and gradually increased in number with age progression. Affymetrix GeneChip array analysis of lung mRNA showed increased levels of proinflammatory cytokine (including IL-1beta, IL-6, and TNF-alpha) and matrix metalloproteinase (including MMP-8, MMP-9, and MMP-12) expression in lal-/- mice. With age progression, some areas of lal-/- mice developed severe abnormal cell proliferation and alveolar remodeling. In other areas, alveolar destruction (i.e., emphysema) was observed. In addition, Clara cell hypertrophy and hyperplasia developed in conducting airways. The pathophysiological phenotypes in the lal-/- mouse lungs became more severe with increasing age. The studies support the concept that neutral lipid metabolites play essential roles in pulmonary homeostasis, inflammatory responses, remodeling, and injury repair.
Fat digestion in humans and some mammals such as dogs requires the successive intervention of two lipases: gastric lipase, which is stable and active despite the highly acidic stomach environment, followed by the classical pancreatic lipase secreted into the duodenum. We previously solved the structure of recombinant human gastric lipase (HGL) at 3.0-A resolution in its closed form; this was the first structure to be described within the mammalian acid lipase family. Here we report on the open structure of the recombinant dog gastric lipase (r-DGL) at 2.7-A resolution in complex with the undecyl-butyl (C11Y4) phosphonate inhibitor. HGL and r-DGL show 85.7% amino acid sequence identity, which makes it relevant to compare the forms from two different species. The open r-DGL structure confirms the previous description of the HGL catalytic triad (Ser(153), His(353), and Asp(324)) with the catalytic serine buried and an oxyanion hole (NH groups of Gln(154) and Leu(67)). In r-DGL, the binding of the C11Y4 phosphonate inhibitor induces part of the cap domain, the lid, to roll over the enzyme surface and to expose a catalytic crevice measuring approximately 20 x 20 x 7 A(3). The C11Y4 phosphonate fits into this crevice, and a molecule of beta-octyl glucoside fills up the crevice. The C11Y4 phosphonate inhibitor and the detergent molecule suggest a possible binding mode for the natural substrates, the triglyceride molecules.
Fat digestion in humans requires not only the classical pancreatic lipase but also gastric lipase, which is stable and active despite the highly acidic stomach environment. We have solved the structure of recombinant human gastric lipase at 3.0 A resolution, the first structure to be described within the mammalian acid lipase family. This globular enzyme (379 residues) consists of a core domain, belonging to the alpha/beta hydrolase fold family, and an extrusion domain. It possesses a classical catalytic triad (Ser 153, His 353, Asp 324) and an oxyanion hole (NH groups of Gln 154 and Leu 67). Four N-glycosylation sites were identified on the electron density maps. The catalytic serine is deeply buried under the extrusion domain, which is composed of a 'cap' domain and a segment consisting of 30 residues, which can be defined as a lid. Its displacement is necessary for the substrates to access the active site. A phosphonate inhibitor was positioned in the active site which clearly suggests the location of the hydrophobic substrate binding site.
Fat digestion in humans requires not only the classical pancreatic lipase but also gastric lipase, which is stable and active despite the highly acidic stomach environment. We report here the structure of recombinant human gastric lipase at 3.0-A resolution, the first structure to be described within the mammalian acid lipase family. This globular enzyme (379 residues) consists of a core domain belonging to the alpha/beta hydrolase-fold family and a "cap" domain, which is analogous to that present in serine carboxypeptidases. It possesses a classical catalytic triad (Ser-153, His-353, Asp-324) and an oxyanion hole (NH groups of Gln-154 and Leu-67). Four N-glycosylation sites were identified on the electron density maps. The catalytic serine is deeply buried under a segment consisting of 30 residues, which can be defined as a lid and belonging to the cap domain. The displacement of the lid is necessary for the substrates to have access to Ser-153. A phosphonate inhibitor was positioned in the active site that clearly suggests the location of the hydrophobic substrate binding site. The lysosomal acid lipase was modeled by homology, and possible explanations for some previously reported mutations leading to the cholesterol ester storage disease are given based on the present model.
        
Title: Occurrence of a mutation associated with Wolman disease in a family with cholesteryl ester storage disease Maslen CL, Babcock D, Illingworth DR Ref: J Inherit Metab Dis, 18:620, 1995 : PubMed
Cholesteryl ester storage disease (CESD) and Wolman disease (McKusick 278000) are two distinct autosomal recessive disorders, both attributable to a severe reduction in acid cholesteryl ester hydrolase/lysosomal acid lipase activity (EC 3.1.1.13). We have identified compound heterozygous mutations in a family with two siblings affected with CESD. Molecular genetic analysis revealed two mutations one of which has previously been seen only in Wolman disease. Analysis of these mutations acting in concert provides new insight into the correlation of genotype with phenotype in these allelic disorders.
        
Title: Mutations at the lysosomal acid cholesteryl ester hydrolase gene locus in Wolman disease Anderson RA, Byrum RS, Coates PM, Sando GN Ref: Proceedings of the National Academy of Sciences of the United States of America, 91:2718, 1994 : PubMed
The genomic sequences encoding the human lysosomal acid lipase/cholesteryl esterase (sterol esterase; EC 3.1.1.13) have been isolated and sequenced, and the information has been used to identify mutations in both alleles of the gene from a patient with Wolman disease, an autosomal recessive lysosomal lipid storage disorder. The genomic locus consists of 10 exons spread over 36 kb. The 5' flanking region is G+C-rich and has characteristics of a "housekeeping" gene promoter. One of the identified mutations involves the insertion of a T residue after position 634, resulting in the appearance of an in-frame translation stop signal 13 codons downstream. The second mutation is a T-to-C transition at nucleotide 638. This results in a leucine-to-proline substitution at amino acid 179 and is predicted to lead to the disruption of the alpha-helical structure in a highly conserved region of the protein. These mutations are each capable of completely disrupting the catalytic function of the lysosomal acid cholesteryl ester hydrolase; their presence can account for the extreme phenotype of the lysosomal lipid storage disorder manifested in members of this patient's family.