Shayman JA

References (28)

Title : Inhibition of lysosomal phospholipase A2 predicts drug-induced phospholipidosis - Hinkovska-Galcheva_2021_J.Lipid.Res_62_100089
Author(s) : Hinkovska-Galcheva V , Treadwell T , Shillingford JM , Lee A , Abe A , Tesmer JJG , Shayman JA
Ref : J Lipid Res , 62 :100089 , 2021
Abstract : Phospholipidosis, the excessive accumulation of phospholipids within lysosomes, is a pathological response observed following exposure to many drugs across multiple therapeutic groups. A clear mechanistic understanding of the causes and implications of this form of drug toxicity has remained elusive. We previously reported the discovery and characterization of a lysosome-specific phospholipase A2 (PLA2G15) and later reported that amiodarone, a known cause of drug-induced phospholipidosis, inhibits this enzyme. Here, we assayed a library of 163 drugs for inhibition of PLA2G15 to determine whether this phospholipase was the cellular target for therapeutics other than amiodarone that cause phospholipidosis. We observed that 144 compounds inhibited PLA2G15 activity. Thirty-six compounds not previously reported to cause phospholipidosis inhibited PLA2G15 with IC(50) values less than 1 mM and were confirmed to cause phospholipidosis in an in vitro assay. Within this group, fosinopril was the most potent inhibitor (IC(50) 0.18 microM). Additional characterization of the inhibition of PLA2G15 by fosinopril was consistent with interference of PLA2G15 binding to liposomes. PLA2G15 inhibition was more accurate in predicting phospholipidosis compared with in silico models based on pKa and ClogP, measures of protonation, and transport-independent distribution in the lysosome, respectively. In summary, PLA2G15 is a primary target for cationic amphiphilic drugs that cause phospholipidosis, and PLA2G15 inhibition by cationic amphiphilic compounds provides a potentially robust screening platform for potential toxicity during drug development.
ESTHER : Hinkovska-Galcheva_2021_J.Lipid.Res_62_100089
PubMedSearch : Hinkovska-Galcheva_2021_J.Lipid.Res_62_100089
PubMedID: 34087196

Title : Structural Basis of Lysosomal Phospholipase A2 Inhibition by Zn(2) - Bouley_2019_Biochemistry_58_1709
Author(s) : Bouley RA , Hinkovska-Galcheva V , Shayman JA , Tesmer JJG
Ref : Biochemistry , 58 :1709 , 2019
Abstract : Lysosomal phospholipase A2 (LPLA2/PLA2G15) is a key enzyme involved in lipid homeostasis and is characterized by both phospholipase A2 and transacylase activity and by an acidic pH optimum. Divalent cations such as Ca(2+) and Mg(2+) have previously been shown to have little effect on the activity of LPLA2, but the discovery of a novel crystal form of LPLA2 with Zn(2+) bound in the active site suggested a role for this divalent cation in regulating enzyme activity. In this complex, the cation directly coordinates the serine and histidine of the alpha/beta-hydrolase triad and stabilizes a closed conformation. This closed conformation is characterized by an inward shift of the lid loop, which extends over the active site and effectively blocks access to one of its lipid acyl chain binding tracks. Therefore, we hypothesized that Zn(2+) would inhibit LPLA2 activity at a neutral but not acidic pH because histidine would be positively charged at lower pH. Indeed, Zn(2+) was found to inhibit the esterase activity of LPLA2 in a noncompetitive manner exclusively at a neutral pH (between 6.5 and 8.0). Because lysosomes are reservoirs of Zn(2+) in cells, the pH optimum of LPLA2 might allow it to catalyze acyl transfer unimpeded within the organelle. We conjecture that Zn(2+) inhibition of LPLA2 at higher pH maintains a lower activity of the esterase in environments where its activity is not typically required.
ESTHER : Bouley_2019_Biochemistry_58_1709
PubMedSearch : Bouley_2019_Biochemistry_58_1709
PubMedID: 30830753
Gene_locus related to this paper: human-PLA2G15

Title : Lysosomal phospholipase A2 - Shayman_2019_Biochim.Biophys.Acta.Mol.Cell.Biol.Lipids_1864_932
Author(s) : Shayman JA , Tesmer JJG
Ref : Biochimica & Biophysica Acta Molecular & Cellular Biology Lipids , 1864 :932 , 2019
Abstract : Lysosomal phospholipase A2 (PLA2G15) is a ubiquitous enzyme uniquely characterized by a subcellular localization to the lysosome and late endosome. PLA2G15 has an acidic pH optimum, is calcium independent, and acts as a transacylase in the presence of N-acetyl-sphingosine as an acceptor. Recent studies aided by the delineation of the crystal structure of PLA2G15 have clarified further the catalytic mechanism, sn-1 versus sn-2 specificity, and the basis whereby cationic amphiphilic drugs inhibit its activity. PLA2G15 has recently been shown to hydrolyze short chain oxidized phospholipids which access the catalytic site directly based on their aqueous solubility. Studies on the PLA2G15 null mouse suggest a role for the enzyme in the catabolism of pulmonary surfactant. PLA2G15 may also have a role in host defense and in the processing of lipid antigens for presentation by CD1 proteins. This article is part of a Special Issue entitled Novel functions of phospholipase A2 Guest Editors: Makoto Murakami and Gerard Lambeau.
ESTHER : Shayman_2019_Biochim.Biophys.Acta.Mol.Cell.Biol.Lipids_1864_932
PubMedSearch : Shayman_2019_Biochim.Biophys.Acta.Mol.Cell.Biol.Lipids_1864_932
PubMedID: 30077006
Gene_locus related to this paper: human-PLA2G15

Title : Determinants of pH profile and acyl chain selectivity in lysosomal phospholipase A2 - Hinkovska-Galcheva_2018_J.Lipid.Res_59_1205
Author(s) : Hinkovska-Galcheva V , Kelly R , Manthei KA , Bouley R , Yuan W , Schwendeman A , Tesmer JJG , Shayman JA
Ref : J Lipid Res , 59 :1205 , 2018
Abstract : Lysosomal phospholipase A2 (LPLA2) is characterized by broad substrate recognition, peak activity at acidic pH, and the transacylation of lipophilic alcohols, especially N-acetyl-sphingosine. Prior structural analysis of LPLA2 revealed the presence of an atypical acidic residue, Asp13, in the otherwise hydrophobic active site cleft. We hypothesized that Asp13 contributed to the pH profile and/or substrate preference of LPLA2 for unsaturated acyl chains. To test this hypothesis, we substituted Asp13 for alanine, cysteine, or phenylalanine; then, we monitored the formation of 1-O-acyl-N-acetylsphingosine to measure the hydrolysis of sn-1 versus sn-2 acyl groups on a variety of glycerophospholipids. Substitutions with Asp13 yielded significant enzyme activity at neutral pH (7.4) and perturbed the selectivity for mono- and double-unsaturated acyl chains. However, this position played no apparent role in selecting for either the acyl acceptor or the head group of the glycerophospholipid. Our modeling indicates that Asp13 and its substitutions contribute to the pH activity profile of LPLA2 and to acyl chain selectivity by forming part of a hydrophobic track occupied by the scissile acyl chain.
ESTHER : Hinkovska-Galcheva_2018_J.Lipid.Res_59_1205
PubMedSearch : Hinkovska-Galcheva_2018_J.Lipid.Res_59_1205
PubMedID: 29724779

Title : A fluorometric assay for lysosomal phospholipase A2 activity using fluorescence-labeled truncated oxidized phospholipid - Abe_2018_Anal.Biochem_549_164
Author(s) : Abe A , Hiraoka M , Shayman JA , Ohguro H
Ref : Analytical Biochemistry , 549 :164 , 2018
Abstract : Lysosomal phospholipase A2 (LPLA2) is a key enzyme involved in the homeostasis of cellular phospholipids. Recently, LPLA2 was reported to preferentially degrade some truncated oxidized phospholipids at the sn-1 position. A commercially available, truncated oxidized phospholipid conjugated with a fluorescent dye, 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphoethanolamine-N-[4-(dipyrrometheneboron difluoride) butanoyl] (PGPE-BODIPY), was used to develop a specific assay for this enzyme. When recombinant mouse LPLA2 was incubated with liposomes consisting of 1,2-O-octadecyl-sn-glycero-3-phosphocholine/PGPE-BODIPY under acidic conditions, PGPE-BODIPY was converted to palmitic acid and a polar BODIPY-product. After phase partitioning by chloroform/methanol, the polar BODIPY-product was recovered in the aqueous phase and identified as 1-lyso-PGPE-BODIPY. The formation of 1-lyso-PGPE-BODIPY was quantitatively determined by fluorescent measurements. The Km and Vmax values of the recombinant LPLA2 for PGPE-BODIPY were 5.64 microM and 20.7 micromol/min/mg protein, respectively. Detectable activity against PGPE-BODIPY was present in LPLA2 deficient mouse sera, but the deacylase activity was completely suppressed by treatment with 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF). AEBSF had no effect on LPLA2 activity. The LPLA2 activity of mouse serum pre-treated with AEBSF was specifically and quantitatively determined by this assay method. The PGPE-BODIPY and AEBSF based LPLA2 assay is convenient and can be used to measure LPLA2 activity in a variety of biological specimens.
ESTHER : Abe_2018_Anal.Biochem_549_164
PubMedSearch : Abe_2018_Anal.Biochem_549_164
PubMedID: 29605449

Title : A retractable lid in lecithin:cholesterol acyltransferase provides a structural mechanism for activation by apolipoprotein A-I - Manthei_2017_J.Biol.Chem_292_20313
Author(s) : Manthei KA , Ahn J , Glukhova A , Yuan W , Larkin C , Manett TD , Chang L , Shayman JA , Axley MJ , Schwendeman A , Tesmer JJG
Ref : Journal of Biological Chemistry , 292 :20313 , 2017
Abstract : Lecithin:cholesterol acyltransferase (LCAT) plays a key role in reverse cholesterol transport by transferring an acyl group from phosphatidylcholine to cholesterol, promoting the maturation of high-density lipoproteins (HDL) from discoidal to spherical particles. LCAT is activated through an unknown mechanism by apolipoprotein A-I (apoA-I) and other mimetic peptides that form a belt around HDL. Here, we report the crystal structure of LCAT with an extended lid that blocks access to the active site, consistent with an inactive conformation. Residues Thr-123 and Phe-382 in the catalytic domain form a latch-like interaction with hydrophobic residues in the lid. Because these residues are mutated in genetic disease, lid displacement was hypothesized to be an important feature of apoA-I activation. Functional studies of site-directed mutants revealed that loss of latch interactions or the entire lid enhanced activity against soluble ester substrates, and hydrogen-deuterium exchange (HDX) mass spectrometry revealed that the LCAT lid is extremely dynamic in solution. Upon addition of a covalent inhibitor that mimics one of the reaction intermediates, there is an overall decrease in HDX in the lid and adjacent regions of the protein, consistent with ordering. These data suggest a model wherein the active site of LCAT is shielded from soluble substrates by a dynamic lid until it interacts with HDL to allow transesterification to proceed.
ESTHER : Manthei_2017_J.Biol.Chem_292_20313
PubMedSearch : Manthei_2017_J.Biol.Chem_292_20313
PubMedID: 29030428
Gene_locus related to this paper: human-LCAT

Title : Prolonged Ocular Inflammation in Endotoxin-Induced Uveitis in Lysosomal Phospholipase A2-Deficient Mice - Sawada_2017_Curr.Eye.Res_42_611
Author(s) : Sawada K , Hiraoka M , Abe A , Kelly R , Shayman JA , Ohguro H
Ref : Current Eye Research , 42 :611 , 2017
Abstract : PURPOSE: The goal of present study was to elucidate the pathophysiological roles of lysosomal phospholipase A2 (LPLA2) in intraocular pressure (IOP) levels and ocular inflammation. METHODS: C57BL/6 (wild-type) and LPLA2-deficient mice with C57BL/6 background were employed. The IOPs were compared between wild-type and LPLA2-deficient mice during their aging, after topical administration of antiglaucoma medications such as travoprost, dorzolamide, or timolol maleate, or after induction of endotoxin-induced uveitis (EIU) using lipopolysaccharide (LPS). Concerning the EIU, ocular inflammation was also evaluated by immunohistochemical analysis by the anti-glial fibrillary acidic protein (GFAP) antibody. RESULTS: The LPLA2-deficient mice showed higher IOP levels than the wild-type mice until 2 months of age (P = 1.60E-06); in older mice there was no difference between the two groups. Significant differences in the IOP changes between groups in young mice were seen after administration of 0.5% timolol (P < 0.05). Upon induction of EIU by LPS, compared with wild-type mice (P < 0.05), IOPs were significantly elevated in LPLA2-deficient mice at maximum levels of the ocular inflammation (48 h). Immunohistochemical analysis indicated that LPLA2-deficient mice showed more prolonged expression of GFAP at the inner plexiform layer and inner nuclear layer by EIU than that found in the wild-type mice (P < 0.05). CONCLUSIONS: These results confirm that LPLA2 plays a significant role in the control of IOP during mouse ocular development or with ocular inflammation by facilitating the digestion of intraocular insoluble materials.
ESTHER : Sawada_2017_Curr.Eye.Res_42_611
PubMedSearch : Sawada_2017_Curr.Eye.Res_42_611
PubMedID: 27612621

Title : Preferential hydrolysis of truncated oxidized glycerophospholipids by lysosomal phospholipase A2 - Abe_2017_J.Lipid.Res_58_339
Author(s) : Abe A , Hiraoka M , Ohguro H , Tesmer JJ , Shayman JA
Ref : J Lipid Res , 58 :339 , 2017
Abstract : Truncated oxidized glycerophospholipids (ox-PLs) are bioactive lipids resulting from oxidative stress. The catabolic pathways for truncated ox-PLs are not fully understood. Lysosomal phospholipase A2 (LPLA2) with phospholipase A and transacylase activities is a key enzyme in phospholipid homeostasis. The present study assessed whether LPLA2 could hydrolyze truncated ox-PLs. Incubation of LPLA2 with liposomes consisting of 1,2-O-octadecenyl-sn-glycero-3-phosphocholine (DODPC)/1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) or truncated oxidized phosphatidylcholine (ox-PC)/N-acetylsphingosine (NAS) under acidic conditions resulted in the preferential deacylation at the sn-1 position of the truncated ox-PCs. Additionally, the release of free fatty acid from the truncated ox-PCs preferentially occurred compared with the NAS-acylation. Incubation of LPLA2 with the liposomes consisting of DODPC/DOPC/truncated ox-PC/NAS resulted in the same preferential fatty acid release from the truncated ox-PC. The cationic amphiphilic drug, amiodarone, did not inhibit such fatty acid release, indicating that truncated ox-PCs partition from the lipid membrane into the aqueous phase and react with free LPLA2. Consistent with this mechanism, the hydrolysis of some truncated ox-PCs, but not DOPC, by LPLA2 was detected at neutral pH. Additionally, LPLA2-overexpressed Chinese hamster ovary cells efficiently catabolized truncated ox-PC and were protected from growth inhibition. These findings support the existence of a novel catabolic pathway for truncated ox-PLs via LPLA2.
ESTHER : Abe_2017_J.Lipid.Res_58_339
PubMedSearch : Abe_2017_J.Lipid.Res_58_339
PubMedID: 27993948

Title : Lysosomal Lipases PLRP2 and LPLA2 Process Mycobacterial Multi-acylated Lipids and Generate T Cell Stimulatory Antigens - Gilleron_2016_Cell.Chem.Biol_23_1147
Author(s) : Gilleron M , Lepore M , Layre E , Cala-De Paepe D , Mebarek N , Shayman JA , Canaan S , Mori L , Carriere F , Puzo G , De Libero G
Ref : Cell Chemical Biology , 23 :1147 , 2016
Abstract : Complex antigens require processing within antigen-presenting cells (APCs) to form T cell stimulatory complexes with CD1 antigen-presenting molecules. It remains unknown whether lipids with multi-acylated moieties also necessitate digestion by lipases to become capable of binding CD1 molecules and stimulate T cells. Here, we show that the mycobacterial tetra-acylated glycolipid antigens phosphatidyl-myo-inositol mannosides (PIM) are digested to di-acylated forms by pancreatic lipase-related protein 2 (PLRP2) and lysosomal phospholipase A2 (LPLA2) within APCs. Recombinant PLRP2 and LPLA2 removed the sn1- and sn2-bound fatty acids from the PIM glycerol moiety, as revealed by mass spectrometry and nuclear magnetic resonance studies. PLRP2 or LPLA2 gene silencing in APCs abolished PIM presentation to T cells, thus revealing an essential role of both lipases in vivo. These findings show that endosomal lipases participate in lipid antigen presentation by processing lipid antigens and have a role in T cell immunity against mycobacteria.
ESTHER : Gilleron_2016_Cell.Chem.Biol_23_1147
PubMedSearch : Gilleron_2016_Cell.Chem.Biol_23_1147
PubMedID: 27662254

Title : Structure and function of lysosomal phospholipase A2 and lecithin:cholesterol acyltransferase - Glukhova_2015_Nat.Commun_6_6250
Author(s) : Glukhova A , Hinkovska-Galcheva V , Kelly R , Abe A , Shayman JA , Tesmer JJ
Ref : Nat Commun , 6 :6250 , 2015
Abstract : Lysosomal phospholipase A2 (LPLA2) and lecithin:cholesterol acyltransferase (LCAT) belong to a structurally uncharacterized family of key lipid-metabolizing enzymes responsible for lung surfactant catabolism and for reverse cholesterol transport, respectively. Whereas LPLA2 is predicted to underlie the development of drug-induced phospholipidosis, somatic mutations in LCAT cause fish eye disease and familial LCAT deficiency. Here we describe several high-resolution crystal structures of human LPLA2 and a low-resolution structure of LCAT that confirms its close structural relationship to LPLA2. Insertions in the alpha/beta hydrolase core of LPLA2 form domains that are responsible for membrane interaction and binding the acyl chains and head groups of phospholipid substrates. The LCAT structure suggests the molecular basis underlying human disease for most of the known LCAT missense mutations, and paves the way for rational development of new therapeutics to treat LCAT deficiency, atherosclerosis and acute coronary syndrome.
ESTHER : Glukhova_2015_Nat.Commun_6_6250
PubMedSearch : Glukhova_2015_Nat.Commun_6_6250
PubMedID: 25727495
Gene_locus related to this paper: human-LCAT

Title : Lysosomal phospholipase A2: a novel player in host immunity to Mycobacterium tuberculosis - Schneider_2014_Eur.J.Immunol_44_2394
Author(s) : Schneider BE , Behrends J , Hagens K , Harmel N , Shayman JA , Schaible UE
Ref : European Journal of Immunology , 44 :2394 , 2014
Abstract : Phospholipases catalyze the cleavage of membrane phospholipids into smaller bioactive molecules. The lysosomal phospholipase A2 (LPLA2 ) is specifically expressed in macrophages. LPLA2 gene deletion in mice causes lysosomal phospholipid accumulation in tissue macrophages leading to phospholipidosis. This phenotype becomes most prominent in alveolar macrophages where LPLA2 contributes to surfactant phospholipid degradation. High expression of LPLA2 in alveolar macrophages prompted us to investigate its role in host immunity against the respiratory pathogen Mycobacterium tuberculosis, the causative agent of tuberculosis. Here we report that adaptive immune responses to M. tuberculosis were impaired in LPLA2 deficient mice. Upon aerosol infection with M. tuberculosis, LPLA2 deficient mice showed enhanced mycobacterial counts but less lung immunopathology and pulmonary inflammatory responses. Compromised T-cell priming in the lymph nodes was associated with impaired pulmonary T-cell recruitment and activation. Together with reduced Th1 type cytokine production, these results indicate that LPLA2 is indispensable for the induction of adaptive T-cell immunity to M. tuberculosis. Taken together, we identified an unexpected and novel function of a lysosomal phospholipid-degrading enzyme.
ESTHER : Schneider_2014_Eur.J.Immunol_44_2394
PubMedSearch : Schneider_2014_Eur.J.Immunol_44_2394
PubMedID: 24825529

Title : A fluorogenic phospholipid for the detection of lysosomal phospholipase A2 activity - Abe_2013_Anal.Biochem_434_78
Author(s) : Abe A , Rzepecki PW , Shayman JA
Ref : Analytical Biochemistry , 434 :78 , 2013
Abstract : Lysosomal phospholipase A2 (group XV PLA2, LPLA2) is a lysosomal enzyme linked to drug-induced phospholipidosis. We developed phospholipid "smart probes" based on the conversion of a quenched fluorogenic substrate to a fluorescent product. Due to the preference of LPLA2 for phosphatidylglycerol, three fluorogenic phosphatidylglycerols were synthesized. Two fluorogenic phosphatidylglycerols were conjugated with one FAM (fluorescein amidite) group and one DABCYL [4-(4-dimethylaminophenylazo)-benzoyl] group; the third substrate consisted of two FAM groups conjugated at the sn-1 and sn-2 positions. The sn-1 ester linkage was replaced with an amide linkage. 1-FAM-2-DABCYL-PG was degraded by recombinant LPLA2 and mouse serum but not by the serum obtained from LPLA2-deficient mice when 1,2-dioleoyl-PG/1-FAM-2-DABCYL-PG liposomes were used. The formation of 1-FAM-lyso-PG generated from 1-FAM-2-DABCYL-PG in the presence of LPLA2 was quantitatively determined by fluorescent measurements. The 1-FAM-2-DABCYL-PG incorporated into 1,2-dioleoyl-phosphatidylcholine/sulfatide liposomes was used to evaluate the effect of the cationic amphiphilic drugs amiodarone and fluoxetine on LPLA2 activity. The IC(50) values of amiodarone and fluoxetine estimated by fluorescent measurement were 10 and 19microM, respectively. These results indicate that 1-FAM-2-DABCYL-PG is a specific substrate for LPLA2 and a useful reagent for the detection of LPLA2 activity from multiple sources.
ESTHER : Abe_2013_Anal.Biochem_434_78
PubMedSearch : Abe_2013_Anal.Biochem_434_78
PubMedID: 23146589

Title : Role for lysosomal phospholipase A2 in iNKT cell-mediated CD1d recognition - Paduraru_2013_Proc.Natl.Acad.Sci.U.S.A_110_5097
Author(s) : Paduraru C , Bezbradica JS , Kunte A , Kelly R , Shayman JA , Veerapen N , Cox LR , Besra GS , Cresswell P
Ref : Proc Natl Acad Sci U S A , 110 :5097 , 2013
Abstract : Invariant natural killer T (iNKT) cells recognize self lipid antigens presented by CD1d molecules. The nature of the self-antigens involved in the development and maturation of iNKT cells is poorly defined. Lysophospholipids are self-antigens presented by CD1d that are generated through the action of phospholipases A1 and A2. Lysosomal phospholipase A2 (LPLA2, group XV phospholipase A2) resides in the endocytic system, the main site where CD1d antigen acquisition occurs, suggesting that it could be particularly important in CD1d function. We find that Lpla2(-/-) mice show a decrease in iNKT cell numbers that is neither the result of a general effect on the development of lymphocyte populations nor of effects on CD1d expression. However, endogenous lipid antigen presentation by CD1d is reduced in the absence of LPLA2. Our data suggest that LPLA2 plays a role in the generation of CD1d complexes with thymic lipids required for the normal selection and maturation of iNKT cells.
ESTHER : Paduraru_2013_Proc.Natl.Acad.Sci.U.S.A_110_5097
PubMedSearch : Paduraru_2013_Proc.Natl.Acad.Sci.U.S.A_110_5097
PubMedID: 23493550

Title : 1-O-acylceramides are natural components of human and mouse epidermis - Rabionet_2013_J.Lipid.Res_54_3312
Author(s) : Rabionet M , Bayerle A , Marsching C , Jennemann R , Grone HJ , Yildiz Y , Wachten D , Shaw W , Shayman JA , Sandhoff R
Ref : J Lipid Res , 54 :3312 , 2013
Abstract : The lipid-rich stratum corneum functions as a barrier against pathogens and desiccation inter alia by an unbroken meshwork of extracellular lipid lamellae. These lamellae are composed of cholesterol, fatty acids, and ceramides (Cers) in an equimolar ratio. The huge class of skin Cers consists of three groups: group I, "classical" long and very long chain Cers; group II, ultra-long chain Cers; and group III, omega-esterified ultra-long chain Cers, which are esterified either with linoleic acid or with cornified envelope proteins and are required for the water permeability barrier. Here, we describe 1-O-acylceramides as a new class of epidermal Cers in humans and mice. These Cers contain, in both the N- and 1-O-position, long to very long acyl chains. They derive from the group I of classical Cers and make up 5% of all esterified Cers. Considering their chemical structure and hydrophobicity, we presume 1-O-acylceramides to contribute to the water barrier homeostasis. Biosynthesis of 1-O-acylceramides is not dependent on lysosomal phospholipase A2. However, glucosylceramide synthase deficiency was followed by a 7-fold increase of 1-O-acylceramides, which then contributed 30% to all esterified Cers. Furthermore, loss of neutral glucosylceramidase resulted in decreased levels of a 1-O-acylceramide subgroup. Therefore, we propose 1-O-acylceramides to be synthesized at endoplasmic reticulum-related sites.
ESTHER : Rabionet_2013_J.Lipid.Res_54_3312
PubMedSearch : Rabionet_2013_J.Lipid.Res_54_3312
PubMedID: 24078707

Title : Drug induced phospholipidosis: an acquired lysosomal storage disorder - Shayman_2013_Biochim.Biophys.Acta_1831_602
Author(s) : Shayman JA , Abe A
Ref : Biochimica & Biophysica Acta , 1831 :602 , 2013
Abstract : There is a strong association between lysosome enzyme deficiencies and monogenic disorders resulting in lysosomal storage disease. Of the more than 75 characterized lysosomal proteins, two thirds are directly linked to inherited diseases of metabolism. Only one lysosomal storage disease, Niemann-Pick disease, is associated with impaired phospholipid metabolism. However, other phospholipases are found in the lysosome but remain poorly characterized. A recent exception is lysosomal phospholipase A2 (group XV phospholipase A2). Although no inherited disorder of lysosomal phospholipid metabolism has yet been associated with a loss of function of this lipase, this enzyme may be a target for an acquired form of lysosomal storage, drug induced phospholipidosis. This article is part of a Special Issue entitled Phospholipids and Phospholipid Metabolism.
ESTHER : Shayman_2013_Biochim.Biophys.Acta_1831_602
PubMedSearch : Shayman_2013_Biochim.Biophys.Acta_1831_602
PubMedID: 22960355

Title : Group XV phospholipase A, a lysosomal phospholipase A - Shayman_2011_Prog.Lipid.Res_50_1
Author(s) : Shayman JA , Kelly R , Kollmeyer J , He Y , Abe A
Ref : Prog Lipid Res , 50 :1 , 2011
Abstract : A phospholipase A was identified from MDCK cell homogenates with broad specificity toward glycerophospholipids including phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and phosphatidylglycerol. The phospholipase has the unique ability to transacylate short chain ceramides. This phospholipase is calcium-independent, localized to lysosomes, and has an acidic pH optimum. The enzyme was purified from bovine brain and found to be a water-soluble glycoprotein consisting of a single peptide chain with a molecular weight of 45 kDa. The primary structure deduced from the DNA sequences is highly conserved between chordates. The enzyme was named lysosomal phospholipase A (LPLA) and subsequently designated group XV phospholipase A. LPLA has 49% of amino acid sequence identity to lecithin-cholesterol acyltransferase and is a member of the alphabeta-hydrolase superfamily. LPLA is highly expressed in alveolar macrophages. A marked accumulation of glycerophospholipids and extensive lamellar inclusion bodies, a hallmark of cellular phospholipidosis, is observed in alveolar macrophages in LPLA(-/-) mice. This defect can also be reproduced in macrophages that are exposed to cationic amphiphilic drugs such as amiodarone. In addition, older LPLA(-/-) mice develop a phenotype similar to human autoimmune disease. These observations indicate that LPLA may play a primary role in phospholipid homeostasis, drug toxicity, and host defense.
ESTHER : Shayman_2011_Prog.Lipid.Res_50_1
PubMedSearch : Shayman_2011_Prog.Lipid.Res_50_1
PubMedID: 21074554
Gene_locus related to this paper: human-PLA2G15

Title : The measurement of lysosomal phospholipase A2 activity in plasma - Abe_2010_J.Lipid.Res_51_2464
Author(s) : Abe A , Kelly R , Shayman JA
Ref : J Lipid Res , 51 :2464 , 2010
Abstract : A deficiency of lysosomal phospholipase A2 (LPLA2) causes macrophage-associated phospholipidosis, suggesting that the enzyme is important in the lipid catabolism. Because LPLA2 is secreted by macrophages, extracellular LPLA2 activity may potentially reflect a change in macrophage activation. In this report, the detection of LPLA2 activity in plasma was established by the measurement of the transacylase activity of LPLA2 under acidic conditions. No transacylase activity of LPLA2 was detected in normal human plasma when the plasma was incubated with liposomes consisting of 1,2-dioleoylphosphatidylcholine/sulfatide/N-acetylsphingosine (NAS) at pH 4.5. However, the transacylase activity in the plasma was detected when liposomes consisting of 1,2-dioleoylphosphatidylglycerol/NAS were used as a substrate. To establish the specificity of the assay, ceramide transacylase activity was detected in the plasma of wild-type mice. By contrast, the plasma obtained from LPLA2-knockout mice had no measurable transacylase activity under the same conditions. The enzymatic activity of recombinant LPLA2 was inhibited by treatment with methylarachidonylfluorophosphonate. The inhibitor also suppressed the transacylase activity observed in both normal human and wild-type mouse plasma, establishing that the transacylase activity observed in plasma is due to LPLA2. Plasma LPLA2 activity may be a useful bioassay marker for the identification of LPLA2-related disorders.
ESTHER : Abe_2010_J.Lipid.Res_51_2464
PubMedSearch : Abe_2010_J.Lipid.Res_51_2464
PubMedID: 20410020

Title : The role of negatively charged lipids in lysosomal phospholipase A2 function - Abe_2009_J.Lipid.Res_50_2027
Author(s) : Abe A , Shayman JA
Ref : J Lipid Res , 50 :2027 , 2009
Abstract : Lysosomal phospholipase A2 (LPLA2) is characterized by increased activity toward zwitterionic phospholipid liposomes containing negatively charged lipids under acidic conditions. The effect of anionic lipids on LPLA2 activity was investigated. Mouse LPLA2 activity was assayed as C2-ceramide transacylation. Sulfatide incorporated into liposomes enhanced LPLA2 activity under acidic conditions and was weakened by NaCl or increased pH. Amiodarone, a cationic amphiphilic drug, reduced LPLA2 activity. LPLA2 exhibited esterase activity when p-nitro-phenylbutyrate (pNPB) was used as a substrate. Unlike the phospholipase A2 activity, the esterase activity was detected over wide pH range and not inhibited by NaCl or amiodarone. Presteady-state kinetics using pNPB were consistent with the formation of an acyl-enzyme intermediate. C2-ceramide was an acceptor for the acyl group of the acyl-enzyme but was not available as the acyl group acceptor when dispersed in liposomes containing amiodarone. Cosedimentation of LPLA2 with liposomes was enhanced in the presence of sulfatide and was reduced by raising NaCl, amiodarone, or pH in the reaction mixture. LPLA2 adsorption to negatively charged lipid membrane surfaces through an electrostatic attraction, therefore, enhances LPLA2 enzyme activity toward insoluble substrates. Thus, anionic lipids present within lipid membranes enhance the rate of phospholipid hydrolysis by LPLA2 at lipid-water interfaces.
ESTHER : Abe_2009_J.Lipid.Res_50_2027
PubMedSearch : Abe_2009_J.Lipid.Res_50_2027
PubMedID: 19321879

Title : The secretion and uptake of lysosomal phospholipase A2 by alveolar macrophages - Abe_2008_J.Immunol_181_7873
Author(s) : Abe A , Kelly R , Kollmeyer J , Hiraoka M , Lu Y , Shayman JA
Ref : J Immunol , 181 :7873 , 2008
Abstract : Macrophages have long been known to secrete a Phospholipase A(2) with an acidic pH optimum in response to phagocytic stimuli. However, the enzyme or enzymes responsible for this activity have not been identified. We report that mouse alveolar macrophages release lysosomal phospholipase A(2) (LPLA(2)) into the medium of cultured cells following stimulation with zymosan. The release of the enzyme was detected by enzymatic activity assays as well as by Western blotting using an Ab against mouse LPLA(2). LPLA(2) is a high mannose type glycoprotein found in lysosomes, suggesting that the released enzyme might be reincorporated into alveolar macrophages via a mannose or mannose phosphate receptor. Recombinant glycosylated mouse LPLA(2) produced by HEK293 cells was applied to LPLA(2)-deficient (LPLA(2)(-/-)) mouse alveolar macrophages. The uptake of exogenous LPLA(2) into LPLA(2)(-/-) alveolar macrophages occurred in a concentration-dependent manner. The LPLA(2) taken into the alveolar macrophages colocalized with the lysosomal marker, Lamp-1. This uptake was significantly suppressed in the presence of alpha-methyl-mannoside but not in the presence of mannose 6-phosphate. Thus, the predominant pathway for uptake of exogenous LPLA(2) is via the mannose receptor, with subsequent translocation into acidic, Lamp-1-associated compartments. LPLA(2)(-/-) alveolar macrophages are characterized by marked accumulation of phosphatidylcholine and phosphatidylethanolamine. Treatment with the recombinant LPLA(2) rescued the LPLA(2)(-/-) alveolar macrophages by markedly decreasing the phospholipid accumulation. The application of a catalytically inactive LPLA(2) revealed that the enzymatic activity of LPLA(2) was required for the phospholipid reduction. These studies identify LPLA(2) as a high m.w.-secreted Phospholipase A(2).
ESTHER : Abe_2008_J.Immunol_181_7873
PubMedSearch : Abe_2008_J.Immunol_181_7873
PubMedID: 19017977

Title : The acylation of lipophilic alcohols by lysosomal phospholipase A2 - Abe_2007_J.Lipid.Res_48_2255
Author(s) : Abe A , Hiraoka M , Shayman JA
Ref : J Lipid Res , 48 :2255 , 2007
Abstract : A novel lysosomal phospholipase A(2) (LPLA2) with specificity toward phosphatidylethanolamine and phosphatidylcholine was previously purified and cloned. LPLA2 transfers sn-1 or sn-2 acyl groups of phospholipids to the C1 hydroxyl of the short-chain ceramide N-acetylsphingosine (NAS) under acidic conditions. The common features of lipophilic alcohols serving as acceptor molecules in the transacylase reaction were examined. 1-O-Hexadecyl-2-acetyl-sn-glycerol (HAG) was acylated by LPLA2 similar to NAS. HAG competed with NAS and inhibited NAS acylation. The transacylation of 1-O-hexadecyl-glycerol (HG), 1-O-palmityl-2-O-methyl-sn-glycerol (PMG), and monoacylglycerols was also investigated. HG, PMG, 1- or 3-palmitoyl-sn-glycerol, and 2-palmitoylglycerol were converted to 1,3-alkylacylglycerol, 1,2-dialkyl-3-acylglycerol, 1,3-diacylglycerol, and 1,2- or 2,3-diacylglycerol, respectively. HG and monoacylglycerol inhibited the acylation of NAS by the enzyme with IC(50) values of 35 and 45 microM, respectively. Additionally, the enzyme acylated glycerol to produce 1- or 3-acyl-sn-glycerol but not 2-acylglycerol. Therefore, the preferred acceptor molecules for LPLA2 are primary alcohols with one long carbon chain and one small nonpolar residue linked to the C2 position of ethanol. The enzyme acylated other natural lipophilic alcohols, including anandamide and oleoylethanolamide. Thus, LPLA2 may function to remodel acyl groups and modulate the biological and pharmacological activities of some lipophilic alcohols.
ESTHER : Abe_2007_J.Lipid.Res_48_2255
PubMedSearch : Abe_2007_J.Lipid.Res_48_2255
PubMedID: 17626977

Title : A role for lysosomal phospholipase A2 in drug induced phospholipidosis - Abe_2007_Drug.Metab.Lett_1_49
Author(s) : Abe A , Hiraoka M , Shayman JA
Ref : Drug Metab Lett , 1 :49 , 2007
Abstract : Many therapeutic drugs currently in use are cationic amphiphiles. These cationic amphiphilic drugs (CADs) induce phospholipidosis in humans and experimental animals. The recent study shows that CAD-induced cellular phospholipidosis is linked to the impairment of phospholipid catabolism by inhibition of lysosomal phospholipase A2 activity.
ESTHER : Abe_2007_Drug.Metab.Lett_1_49
PubMedSearch : Abe_2007_Drug.Metab.Lett_1_49
PubMedID: 19356018

Title : Lysosomal phospholipase A2 and phospholipidosis - Hiraoka_2006_Mol.Cell.Biol_26_6139
Author(s) : Hiraoka M , Abe A , Lu Y , Yang K , Han X , Gross RW , Shayman JA
Ref : Molecular & Cellular Biology , 26 :6139 , 2006
Abstract : A lysosomal phospholipase A2, LPLA2, was recently characterized and shown to have substrate specificity for phosphatidylcholine and phosphatidylethanolamine. LPLA2 is ubiquitously expressed but is most highly expressed in alveolar macrophages. Double conditional gene targeting was employed to elucidate the function of LPLA2. LPLA2-deficient mice (Lpla2-/-) were generated by the systemic deletion of exon 5 of the Lpla2 gene, which encodes the lipase motif essential for the phospholipase A2 activity. The survival of the Lpla2-/- mice was normal. Lpla2-/- mouse mating pairs yielded normal litter sizes, indicating that the gene deficiency did not impair fertility or fecundity. Alveolar macrophages from wild-type but not Lpla2-/- mice readily degraded radiolabeled phosphatidylcholine. A marked accumulation of phospholipids, in particular phosphatidylethanolamine and phosphatidylcholine, was found in the alveolar macrophages, the peritoneal macrophages, and the spleens of Lpla2-/- mice. By 1 year of age, Lpla2-/- mice demonstrated marked splenomegaly and increased lung surfactant phospholipid levels. Ultrastructural examination of Lpla2-/- mouse alveolar and peritoneal macrophages revealed the appearance of foam cells with lamellar inclusion bodies, a hallmark of cellular phospholipidosis. Thus, a deficiency of lysosomal phospholipase A2 results in foam cell formation, surfactant lipid accumulation, splenomegaly, and phospholipidosis in mice.
ESTHER : Hiraoka_2006_Mol.Cell.Biol_26_6139
PubMedSearch : Hiraoka_2006_Mol.Cell.Biol_26_6139
PubMedID: 16880524

Title : Positional specificity of lysosomal phospholipase A2 - Abe_2006_J.Lipid.Res_47_2268
Author(s) : Abe A , Hiraoka M , Shayman JA
Ref : J Lipid Res , 47 :2268 , 2006
Abstract : Lysosomal phospholipase A(2) (Lpla2) is highly expressed in alveolar macrophages and may mediate the phospholipid metabolism of surfactant. Studies on the properties of this phospholipase are consistent with the presence of both phospholipase A(1) and phospholipase A(2) activities. These activities were studied through the production of O-acyl compounds, produced by the transacylase activity of Lpla2. Liposomes containing POPC and N-acetylsphingosine (NAS) were incubated with the soluble fraction obtained from MDCK cells stably transfected with the mouse Lpla2 gene. Two 1-O-acyl-NASs, 1-O-palmitoyl-NAS and 1-O-oleoyl-NAS, were produced by Lpla2. The formation rate of 1-O-oleoyl-NAS was 2.5-fold that of 1-O-palmitoyl-NAS. When 1-oleoyl-2-palmitoyl-sn-glycero-3-phosphocholine (OPPC) was used, the formation rate of 1-O-oleoyl-NAS was 5-fold higher than that of 1-O-palmitoyl-NAS. Thus, Lpla2 can act on acyl groups at both sn-1 and sn-2 positions of POPC and OPPC. When 1-palmitoyl-2-unsaturated acyl-sn-glycero-3-phosphocholines were used as acyl donors, the transacylation of the acyl group from the sn-2 position to NAS was preferred to that of the palmitoyl group from the sn-1 position. An exception was observed for 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PAPC), for which the formation rate of 1-O-palmitoyl-NAS from PAPC was 4-fold greater than that of 1-O-arachidonoyl-NAS. Thus, Lpla2 has broad positional specificity for the sn-1 and sn-2 acyl groups in phosphatidylcholine and phosphatidylethanolamine.
ESTHER : Abe_2006_J.Lipid.Res_47_2268
PubMedSearch : Abe_2006_J.Lipid.Res_47_2268
PubMedID: 16837646

Title : Structure and function of lysosomal phospholipase A2: identification of the catalytic triad and the role of cysteine residues - Hiraoka_2005_J.Lipid.Res_46_2441
Author(s) : Hiraoka M , Abe A , Shayman JA
Ref : J Lipid Res , 46 :2441 , 2005
Abstract : Lysosomal phospholipase A2 (LPLA2) is an acidic phospholipase that is highly expressed in alveolar macrophages and that may play a role in the catabolism of pulmonary surfactant. The primary structure found in LCAT is conserved in LPLA2, including three amino acid residues potentially required for catalytic activity and four cysteine residues. LPLA2 activity was measured in COS-7 cells transfected with c-myc-conjugated mouse LPLA2 (mLPLA2) or mutated LPLA2. Single alanine substitutions in the catalytic triad resulted in the elimination of LPLA2 activity. Four cysteine residues (C65, C89, C330, and C371), conserved between LPLA2 and LCAT, were replaced with alanine. Quadruple mutations at C65, C89, C330, and C371, double mutations at C65 and C89, and a single mutation at C65 or C89 resulted in the elimination of activity. Double mutations at C330 and C371 and a single mutation at C330 or C371 resulted in a partial reduction of activity. Thus, the presence of a disulfide bond between C330 and C371 is not required for LPLA2 activity. We propose that one disulfide bond between C65 and C89 and free cysteine residues at C330 and C371 and the triad, serine-198, aspartic acid-360, and histidine-392, are required for the full expression of mLPLA2 activity.
ESTHER : Hiraoka_2005_J.Lipid.Res_46_2441
PubMedSearch : Hiraoka_2005_J.Lipid.Res_46_2441
PubMedID: 16106046

Title : Induction of lysosomal phospholipase A2 through the retinoid X receptor in THP-1 cells - Abe_2004_J.Lipid.Res_45_667
Author(s) : Abe A , Poucher HK , Hiraoka M , Shayman JA
Ref : J Lipid Res , 45 :667 , 2004
Abstract : An acidic phospholipase A(2) (LPLA(2)) was recently purified and cloned. THP-1 cells were used to characterize the gene induction of LPLA(2). THP-1 cells were stimulated with several differentiation agents. The LPLA(2) mRNA and activity increased in cells treated with phorbol ester but not with vitamin D3, interferon-gamma, or granulocyte macrophage colony-stimulating factor. All-trans-retinoic acid enhanced mRNA expression and enzyme activity in a dose- and time-dependent manner. The natural 9-cis and 13-cis isomers of retinoic acid enhanced transcription and activity. Two classes of nuclear receptors, the retinoic acid receptor (RAR) and the retinoid X receptor (RXR), mediate retinoic acid signaling. Specific RAR and RXR agonists were used to identify the nuclear receptor responsible for LPLA(2) induction by retinoic acid. Treatment with the RAR agonist 4-[E-2-tetrahydro-5,5,8,8-tetra-methyl-2-naphthalenyl]1-propenyl benzoic acid (TTNPB) resulted in a small and statistically significant increase of the mRNA expression and activity of LPLA(2). The RXR agonist methoprene acid worked as well as all-trans-retinoic acid at increasing both mRNA and enzyme activity. The methoprene acid and TTNPB effects were not synergistic. The peroxisome proliferator-activated receptor gamma agonists 15-deoxy-Delta(12,14)-prostaglandin J(2) and troglitazone failed to induce LPLA(2) activity and mRNA. Thus, an RXR-dependent pathway controls LPLA(2) gene activation by retinoic acid in THP-1 cells.
ESTHER : Abe_2004_J.Lipid.Res_45_667
PubMedSearch : Abe_2004_J.Lipid.Res_45_667
PubMedID: 14754907

Title : Lysosomal phospholipase A2 is selectively expressed in alveolar macrophages - Abe_2004_J.Biol.Chem_279_42605
Author(s) : Abe A , Hiraoka M , Wild S , Wilcoxen SE , Paine R, 3rd , Shayman JA
Ref : Journal of Biological Chemistry , 279 :42605 , 2004
Abstract : Lung surfactant is the surface-active agent comprised of phospholipids and proteins that lines pulmonary alveoli. Surfactant stabilizes the alveolar volume by reducing surface tension. Previously, we identified a lysosomal phospholipase A2, termed LPLA2, with specificity toward phosphatidylcholine and phosphatidylethanolamine. The phospholipase is localized to lysosomes, is calcium-independent, has an acidic pH optimum, and transacylates ceramide. Here, we demonstrate that LPLA2 is selectively expressed in alveolar macrophages but not in peritoneal macrophages, peripheral blood monocytes, or other tissues. Other macrophage-associated phospholipase A2s do not show a comparable distribution. LPLA2 is of high specific activity and recognizes disaturated phosphatidylcholine as a substrate. The lysosomal phospholipase A2 activity is six times lower in alveolar macrophages from mice with a targeted deletion of the granulocyte macrophage colony-stimulating factor (GM-CSF), a model of impaired surfactant catabolism, compared with those from wild-type mice. However, LPLA2 activity and protein levels are measured in GM-CSF null mice in which GM-CSF is expressed as a transgene under the control of the surfactant protein C promoter. Thus LPLA2 may be a major enzyme of pulmonary surfactant phospholipid degradation by alveolar macrophages and may be deficient in disorders of surfactant metabolism.
ESTHER : Abe_2004_J.Biol.Chem_279_42605
PubMedSearch : Abe_2004_J.Biol.Chem_279_42605
PubMedID: 15294901
Gene_locus related to this paper: ratno-q675a5

Title : A turn in the road: How studies on the pharmacology of glucosylceramide synthase inhibitors led to the identification of a lysosomal phospholipase A2 with ceramide transacylase activity - Shayman_2004_Glycoconj.J_20_25
Author(s) : Shayman JA , Abe A , Hiraoka M
Ref : Glycoconj J , 20 :25 , 2004
Abstract : A series of inhibitors of glucosylceramide synthesis, the PDMP based family of compounds, has been developed as a tool for the study of sphingolipid biochemistry and biology. During the course of developing more active glucosylceramide synthase inhibitors, we identified a second site of inhibitory activity for PDMP and its structural homologues that accounted for the ability of the inhibitors to raise cell and tissue ceramide levels. This inhibitory activity was directed against a previously unknown pathway for ceramide metabolism, viz. the formation of 1- O -acylceramide. In this pathway the addition of a fatty acyl group to the primary hydroxyl of ceramide occurs through a transacylation with either phosphatidylethanolamine or phosphatidylcholine as a substrate. However, both in the absence and presence of ceramide, water serves as an acceptor for the fatty acid. Thus the enzyme may be considered to be a phospholipase A2. The enzyme is unique in that it has an acidic pH optimum and is localized to lysosomes by cell fractionation. More recently, the 1- O -acylceramide synthase has been purified, sequenced, and cloned. This phospholipase A2 was discovered to be structurally homologous to lecithin cholesterol acyltransferase (LCAT). However, this phospholipase A2 does not recognize cholesterol and lacks the defined lipoprotein-binding domain present in LCAT. We now refer to this enzyme as lysosomal phospholipase A2 (LPLA2). Although acidic phospholipase A2 activities have been previously identified, LPLA2 appears to be the first lysosomal PLA2 to have been sequenced. This new phospholipase A2 lacks an obvious and proven biological function.
ESTHER : Shayman_2004_Glycoconj.J_20_25
PubMedSearch : Shayman_2004_Glycoconj.J_20_25
PubMedID: 14973367

Title : Cloning and characterization of a lysosomal phospholipase A2, 1-O-acylceramide synthase - Hiraoka_2002_J.Biol.Chem_277_10090
Author(s) : Hiraoka M , Abe A , Shayman JA
Ref : Journal of Biological Chemistry , 277 :10090 , 2002
Abstract : Recently, a novel enzyme, 1-O-acylceramide synthase (ACS), was purified and characterized from bovine brain. This enzyme has both calcium-independent phospholipase A(2) and transacylase activities. The discovery of this enzyme led us to propose a new pathway for ceramide metabolism in which the sn-2-acyl group of either phosphatidylethanolamine or phosphatidylcholine is transferred to the 1-hydroxyl group of ceramide. In this study, the partial amino acid sequences from the purified enzyme revealed that the enzyme contains amino acid sequences identical to those of human lecithin:cholesterol acyltransferase-like lysophospholipase (LLPL). The coding sequences of the mouse, bovine, and human genes were obtained from the respective kidney cDNAs by PCR. The open reading frames of LLPL were cloned into pcDNA3 to generate carboxyl-terminally tagged proteins. The expression of mouse LLPL in COS-7 cells demonstrated that transfected cells had higher transacylase and phospholipase A(2) activities than did non-transfected cells. Immunoprecipitation confirmed that LLPL had ACS activity. There were no significant lecithin:cholesterol acyltransferase and lysophospholipase activities in the mouse LLPL-transfected cells under either acidic or neutral conditions. Amino acid sequences from cDNAs of mouse, human, and bovine LLPLs demonstrated a signal peptide cleavage site, one lipase motif (AXSXG), and several N-linked glycosylation sites in each LLPL molecule. The replacement of serine with alanine in the lipase motif of mouse LLPL resulted in elimination of enzyme activity, indicating that the serine residue is part of the catalytic site. Deglycosylation of mouse, human, and bovine LLPLs yielded core proteins with a molecular mass of 42 kDa without change in enzyme activities. LLPL was post-translationally modified by signal peptide cleavage and N-linked glycosylation, and each mature LLPL had the same size core protein. Subcellular fractionation demonstrated that ACS activity co-localized with N-acetylglucosaminidase. Therefore, LLPL encodes a novel lysosomal enzyme, ACS.
ESTHER : Hiraoka_2002_J.Biol.Chem_277_10090
PubMedSearch : Hiraoka_2002_J.Biol.Chem_277_10090
PubMedID: 11790796
Gene_locus related to this paper: bovin-LPLA2 , canfa-q6xpz3 , human-PLA2G15 , mouse-C87498