Showing papers on "Lipase published in 1992"
TL;DR: The crystal structure of an extracellular triglyceride lipase inhibited irreversibly by diethyl p-nitrophenyl phosphate (E600) was solved by X-ray crystallographic methods and refined to a resolution of 2.65 A.
Abstract: The crystal structure of an extracellular triglyceride lipase (from a fungus Rhizomucor miehei) inhibited irreversibly by diethyl p-nitrophenyl phosphate (E600) was solved by X-ray crystallographic methods and refined to a resolution of 2.65 A. The crystals are isomorphous with those of n-hexylphosphonate ethyl ester/lipase complex [Brzozowski, A. M., Derewenda, U., Derewenda, Z. S., Dodson, G. G., Lawson, D. M., Turkenburg, J. P., Bjorkling, F., Huge-Jensen, B., Patkar, S. A., & Thim, L. (1991) Nature 351, 491-494], where the conformational change was originally observed. The higher resolution of the present study allowed for a detailed analysis of the stereochemistry of the change observed in the inhibited enzyme. The movement of a 15 amino acid long "lid" (residues 82-96) is a hinge-type rigid-body motion which transports some of the atoms of a short alpha-helix (residues 85-91) by over 12 A. There are two hinge regions (residues 83-84 and 91-95) within which pronounced transitions of secondary structure between alpha and beta conformations are caused by dramatic changes of specific conformational dihedral angles (phi and psi). As a result of this change a hydrophobic area of ca. 800 A2 (8% of the total molecule surface) becomes exposed. Other triglyceride lipases are also known to have "lids" similar to the one observed in the R. miehei enzyme, and it is possible that the general stereochemistry of lipase activation at the oil-water interfaces inferred from the present X-ray study is likely to apply to the entire family of lipases.
492 citations
TL;DR: It is proposed that upon lipolytic activation of adipocytes and phosphorylation of HSL by cAMP-dependent protein kinase, the critical event is not an increase in catalytic activity but a translocation of the lipase to its substrate at the surface of the lipid storage droplet.
Abstract: Hormone-sensitive lipase activity (HSL), which is found in the supernatant of centrifuged homogenates of lipolytically quiet isolated rat adipocytes, was greatly reduced in or absent from the supernatant of lipolytically stimulated cells. The lipase was purified 100- to 250-fold from the supernatant of lipolytically quiet cells to 10-20% purity by a single passage over phenyl-Sepharose resin with high (greater than 70%) activity yields. Western blotting of adipocyte homogenate fractions with polyclonal antiserum raised against HSL showed that the enzyme shifted quantitatively from the supernatant of control cells to the floating "fat cake" of lipolytically stimulated cells. A similar shift to the fat cake was observed when cells were disrupted by hypotonic lysis and centrifugation rather than by homogenization. We propose that upon lipolytic activation of adipocytes and phosphorylation of HSL by cAMP-dependent protein kinase, the critical event is not an increase in catalytic activity (i.e., turnover number) but a translocation of the lipase to its substrate at the surface of the lipid storage droplet.
401 citations
TL;DR: The structure of procolipase is described, which essentially consists of three 'fingers' and is topologically comparable to snake toxins and may form the interfacial binding site of pancreatic lipase.
Abstract: Interfacial adsorption of pancreatic lipase is strongly dependent on the physical chemical properties of the lipid surface. These properties are affected by amphiphiles such as phospholipids and bile salts. In the presence of such amphiphiles, lipase binding to the interface requires a protein cofactor, colipase. We obtained crystals of the pancreatic lipase-procolipase complex and solved the structure at 3.04 A resolution. Here we describe the structure of procolipase, which essentially consists of three 'fingers' and is topologically comparable to snake toxins. The tips of the fingers contain most of the hydrophobic amino acids and presumably form the interfacial binding site. Lipase binding occurs at the opposite side to this site and involves polar interactions. Determination of the three-dimensional structure of pancreatic lipase has revealed the presence of two domains: an amino-terminal domain, at residues 1-336 containing the active site and a carboxy-terminal domain at residues 337-449 (ref. 6). Procolipase binds exclusively to the C-terminal domain of lipase. No conformational change in the lipase molecule is induced by the binding of procolipase.
340 citations
TL;DR: The phylogenetic reconstruction suggests that ancestral PL diverged earlier than HL and LPL, which share a more recent root, and a conservation index (CI) is constructed to display conserved structural features within the lipase gene family, a CI of 1.0 signifying perfect conservation.
264 citations
TL;DR: Results presented in this paper strongly suggest that the bile salt-dependent lipase is the only pancreatic enzyme involved in lipid digestion in cod.
229 citations
TL;DR: This study compares two solvents for enzymatic reactions: supercritical carbon dioxide (SCCO2) and organic solvent (n‐hexane) and suggests that activity was in the same range in both media.
Abstract: This study compares two solvents for enzymatic reactions: supercritical carbon dioxide (SCCO2) and organic solvent (n-hexane). The model reaction that was chosen was the esterification of oleic acid by ethanol catalyzed by an immobilized lipase from Mucor miehei (Lypozyme). The stability of the enzyme appeared to be quite good and similar in both media but was affected by the water content. Partition of water between solvents and immobilized enzyme has been calculated from experimental adsorption isotherms. The water content of the solid phase has a dramatic influence on the activity of the enzyme and its optimum value for activity was about 10% (w/w) in both media. A kinetic study enabled a Ping-Pong Bi-Bi reaction mechanism with inhibition by ethanol to be suggested. Despite some differences in kinetic constants, activity was in the same range in both media. Hypotheses for explaining the kinetic constant variations have been proposed and particular attention has been paid to the pH effects.
225 citations
TL;DR: The creation of an artificial interphase between the solvent-immiscible hydrophilic Glycerol and the hydrophobic reaction media by the adsorption of glycerol onto a solid support prior to use was essential for the success of these transformations.
Abstract: Regioisomerically pure 1(3)-rac-monoacylglycerols are conveniently prepared in high yields (>75%) and in multigram quantities by enzymatic esterification of glycerol in the presence of various lipases(Chromobacterium viscosum, Rhizopus delemar, Rhizomucor miehei) with a variety of different acyl donors, such as free fatty acids, fatty acid alkyl esters, vinyl esters and triacylglycerols, as well as natural fats and oils. All reactions are carried out in aprotic organic solvents with low water content, namelyn-hexane, diethyl ether, tBuOMe or mixtures of these solvents. Essential for the success of these transformations were the following two factors. First, the creation of an artificial interphase between the solvent-immiscible hydrophilic glycerol and the hydrophobic reaction medium by its adsorption onto a solid support. Second, a facile system for the separation of the desired monoacylglycerol from the reaction mixture, coupled with the continuous recycling of acyl donor and undesirable by-products.
213 citations
TL;DR: In this paper, the synthesis of peroxytetradecanoic acid from hydrogen peroxide and free carboxylic acid has been investigated using a two phase system of toluene and water.
198 citations
TL;DR: The gene coding for an extracellular lipase of Bacillus subtilis 168 was cloned and found to be expressed in Escherichia coli and revealed a frameshift, due to the presence of an additional adenine in the N-terminal region, which caused the interruption of the open reading frame.
163 citations
TL;DR: The isolated cDNAs coding for two novel human pancreatic lipase-related human proteins, referred to as hPL-related proteins 1 and 2 (hPLRP1 and hPLRP2) and for hPL are shown to show an amino acid sequence identity to hPL, suggesting that the two novel proteins also exist in other species and that some of the sequences reported to be Pancreas lipase might more likely be the orthologues of hPL RP1 or hPL
160 citations
TL;DR: The R- and S-enantiomers of ibuprofen were determined without derivatization by HPLC using a chiral column and the synthesis of esters was profoundly affected by the amount of water in the reaction mixture.
Abstract: Resolution of (R,S)-ibuprofen enantiomers by esterification in different organic solvents was studied using Candida cylindracea lipase. This enzyme preparation had high enantiospecificity for S(+)-ibuprofen in the esterification reaction of a racemic ibuprofen with primary alcohols. The esterification yields of secondary alcohols were much lower than those of primary alcohols. Esterification with tertiary alcohols was not observed. The synthesis of esters was profoundly affected by the amount of water in the reaction mixture. C. cylindracea lipase was active only in very hydrophobic solvents. The esterification activity of the lipase was reduced significantly by addition of water. The R- and S-enantiomers of ibuprofen were determined without derivatization by HPLC using a chiral column.
TL;DR: Major attention is given to molecular studies relating to the regulation of LPL production, to the defects underlying LPL deficiency, and to structure-function relationship of the lipases.
Abstract: Lipoprotein lipase (LPL) is a glycoprotein enzyme that is produced in several cells and tissues. LPL belongs to a large lipase gene family that includes, among others, hepatic lipase and pancreatic lipase. After secretion, LPL becomes anchored on the luminal surface of the capillary endothelial cells. There it hydrolyzes triglycerides in triglyceride-rich lipoproteins, generating free fatty acids that can serve either as a direct energy source or can be stored. Through this action LPL plays a pivotal role both in energy and in lipoprotein metabolism. LPL production is regulated in a tissue-specific fashion by developmental, hormonal, and nutritional factors. The recent availability of the regulatory sequences of the LPL gene will greatly facilitate these regulatory studies in the future. In man, several mutations resulting in familial LPL deficiency have been delineated at a molecular level. The study of these mutations is not only very beneficial from a clinical point of view but also contributes in a major way to our understanding of the structure-function relationship of LPL and other lipases. In this review major attention is given to molecular studies relating to the regulation of LPL production, to the defects underlying LPL deficiency, and to structure-function relationship of the lipases.
TL;DR: The kinetics of reaction are suggested to agree with a ping-pong Bi Bi mechanism in which inhibition by excess of geraniol has been identified.
TL;DR: In this paper, the acyl chain specificity of lipases is evaluated in terms of hydrolysis resistant value (HRV), which is the ratio between the DHA contents in the glyceride mixture of the hydrolyzed oil and original oil.
Abstract: In an attempt to concentrate the content of DHA (docosahexaenoic acid) in a glyceride mixture containing triglyceride, diglyceride and monoglyceride, fish oil was hydrolyzed with six kinds of microbial lipase. After the hydrolysis, free fatty acid was removed and fatty acid components of the glyceride mixtures were analyzed. When the hydrolysis withCandida cylindracea lipase was 70% complete, the DHA content in the glyceride mixture was three times more than that in the original fish oil. The EPA (eicosapentaenoic acid) content became almost 70% of the original fish oil. Hydrolysis with other lipases did not result in an increase in the DHA content in the glyceride mixtures. Hydrolysis of DHA-rich tuna oil (DHA content is about 25%) withCandida cylindracea lipase resulted in 53% DHA in the glyceride mixture. The EPA content, however, remained close to that of the original tuna oil. In this report, the acyl chain specificity of lipases is evaluated in terms of hydrolysis resistant value (HRV). HRV is the ratio between the DHA contents in the glyceride mixture of hydrolyzed oil and original oil. HRV clearly indicates differences in hydrolysis between DHA and other fatty acids (e.g., saturated and monoenoic acids).
TL;DR: In this article, the enantiomeric ratio of porcine pancreatic lipase and Lipase PS in transesterification between trifluoroethyl butanoate and the two unrelated alcohols (±)-sulcatol (1 ) and 3-bromo-5-hydroxymethyl isoxazoline (2 ).
Abstract: The nature of organic solvents markedly influenced the enantioselectivity, expressed as the enantiomeric ratio, of porcine pancreatic lipase and of Lipase PS in transesterification between trifluoroethyl butanoate and the two unrelated alcohols (±)-sulcatol ( 1 ) and (±)-3-bromo-5-hydroxymethyl isoxazoline ( 2 ). However, there was no correlation between enantioselectivity and such physicochemical characteristics of the solvent as hydrophobicity and dielectric constant. A rationale based on the formation of solvent-enzyme complexes is proposed to explain the results. Enzyme enantioselectivity was also affected by temperature but not by the chain length of the acylating agent nor by the removal of water from the reaction medium by molecular sieves. The effects of these parameters on transesterification rates were also investigated.
TL;DR: It is established that the loop with its two amphipathic helices is essential for hydrolysis of long chain fatty acid substrate by LPL, providing new insight into the role of the LPL loop in lipid-substrate interactions.
TL;DR: In this paper, it was shown that the addition of appropriate solid salt hydrates to the reaction mixture is a simple and convenient method to obtain optimal water level conditions throughout the reaction.
TL;DR: These studies support the hypothesis that normal desquamation is mediated by enzymatic modulations in lipid and/or protein content of the SC interstices, and that some forms of pathological or excessive scaling may be due to desmosomal persistence that results from defective or limited delivery of lamellar body‐derived, hydrolytic enzymes to theSC intercellular domains.
Abstract: Summary
Lamellar body secretion results in the delivery of a selected array of hydrolytic enzymes to the extracellular domains of stratum corneum (SC). Deposition and activation of these enzymes in the interstices presumably is associated with the transformation of lamellar body-derived lipids from a relatively polar to a non-polar mixture, as well as the degradation of other non-lipid intercellular substrates. To determine whether abnormal desquamation might result from failure of hydrolytic enzyme delivery to the SC interstices, we localized one catabolic enzyme, acid lipase, previously shown to be a reproducible marker for the lamellar body secretory system, by cytochemical methods within the epidermis of selected human (congenital ichthyosiform erythroderma, CIE) and animal (essential fatty-acid deficient (EFAD) mouse epidermis and mouse tail epidermis) models associated with abnormal scaling or unusual SC retention. In addition, we compared the persistence of desmosomes within normal SC vs. the various models. Normal human and murine epidermis displayed abundant lipase activity both in lamellar bodies (LB) and in association with secreted lamellar body contents in the SC interstices. Despite normal quantities of LB in CIE, EFAD, and mouse tail epidermis, lipase activity was markedly deficient both in LB and in the SC intercellular domains. These studies support the hypothesis that normal desquamation is mediated by enzymatic modulations in lipid and/or protein content of the SC interstices, and that some forms of pathological or excessive scaling may be due to desmosomal persistence that results from defective or limited delivery of lamellar body-derived, hydrolytic enzymes to the SC intercellular domains
TL;DR: It was demonstrated that the stimulation of LPL activity by apolipoprotein C-II and the inhibition of activity by 1 M NaCl originate in structural features within the NH2-terminal domain, and HL is more active than LPL in the hydrolysis of phospholipid substrates.
TL;DR: Information on the catalytic mechanism and on the three-dimensional structure of the P. glumae lipase should facilitate the selection of specific modifications for tailoring this lipase for specific industrial applications.
Abstract: The lipA gene encoding the extracellular lipase produced by Pseudomonas glumae PG1 was cloned and characterized. A sequence analysis revealed an open reading frame of 358 codons encoding the mature lipase (319 amino acids) preceded by a rather long signal sequence of 39 amino acids. As a first step in structure-function analysis, we determined the Ser-Asp-His triad which makes up the catalytic site of this lipase. On the basis of primary sequence homology with other known Pseudomonas lipases, a number of putative active site residues located in conserved areas were found. To determine the residues actually involved in catalysis, we constructed a number of substitution mutants for conserved Ser, Asp, and His residues. These mutant lipases were produced by using P. glumae PG3, from which the wild-type lipase gene was deleted by gene replacement. By following this approach, we showed that Ser-87, Asp-241, and His-285 make up the catalytic triad of the P. glumae lipase. This knowledge, together with information on the catalytic mechanism and on the three-dimensional structure, should facilitate the selection of specific modifications for tailoring this lipase for specific industrial applications.
TL;DR: In this paper, the synthesis of poly(methyl 6-acryloyl-β-galatoside) with M w =196 000 and M n =58 000 for a polydispersity of 2.9 which was highly water soluble and soluble in polar organic solvents.
Abstract: The lipase from Pseudomonas cepacia catalyzed the transesterification of a variety of monosaccharides with vinyl acrylate in pyridine to give the 6-acryloyl esters. The acrylate esters were polymerized in DMF with AIBN to give the poly(acrylate) products. Such a synthetic strategy has led to the synthesis of poly(methyl 6-acryloyl-β-galatoside) with M w =196 000 and M n =58 000 for a polydispersity of 2.9 which was highly water soluble and soluble in polar organic solvents. Addition of ethylene glycol dimethacrylate during AIBN-initiated polymerization in DMF resulted in water insoluble polymrs that swelled in aqueous solutions
TL;DR: A Penicillium simplicissimum strain has been found to produce an inducible extracellular lipase and showed a high stability in water immiscible organic solvents.
TL;DR: Production of extracellular lipase by Rhodotorula glutinis was substantially enhanced when the type and concentration of carbon and nitrogen source, the initial pH of culture medium and the growth temperature were consecutively optimized.
Abstract: Production of extracellular lipase byRhodotorula glutinis was substantially enhanced when the type and concentration of carbon and nitrogen source, the initial pH of culture medium and the growth temperature were consecutively optimized. Lipase activity as high as 30.4 U/ml of culture medium was obtained at optimum conditions, comparing favourably with most of the activities reported for other lipase hyperproducing microorganisms. The enzyme was optimally active at pH 7.5 and 35°C and had, at optimum pH, half-lives of 45 and 11.8 min at 45 and 55°C respectively. The high activity and kinetic characteristics of the enzyme make this process worthy of further investigation.
TL;DR: Two enantiomers of 1,4-dihydropyridine compounds have been obtained with high enantiomeric purity by lipase-catalyzed hydrolysis in organic solvent saturated with water.
01 Jan 1992
TL;DR: The fatty acid specificity of the B-lipase derived from Candida antarctica was investigated in the synthesis of esters of ethyl D-glucopyranoside and lower fatty acids such as hexanoic and octanoic acid and the unsaturated 9-cis-octadecenoic acid were found to be poor substrates of the enzyme.
Abstract: The fatty acid specificity of the B-lipase derived from Candida antarctica was investigated in the synthesis of esters of ethyl D-glucopyranoside. The specificity was almost identical with respect to straight-chain fatty acids with 10 to 18 carbon atoms. However, lower fatty acids such as hexanoic and octanoic acid and the unsaturated 9-cis-octadecenoic acid were found to be poor substrates of the enzyme. As a consequence of this selectivity, these fatty acids were accumulated in the unconverted fraction when ethyl D-glucopyranoside was esterified with an excess of a mixture of fatty acids. This accumulation can reduce the overall effectiveness of the process as the activity of the lipase was found to be reduced when exposed to high concentrations of short-chain fatty acids. Finally, using a simplified experimental set-up, the specificity of the C. antarctica B-lipase was compared to the specificity of lipases derived from C. rugosa, Mucor miehei, Humicola, and Pseudomonas. Apart from the C. rugosa lipase...
TL;DR: Results show that PL responds similarly to carboxylester lipase with respect to changes in interfacial lipid mole fraction in DA-POPC surfaces.(ABSTRACT TRUNCATED at 400 WORDS)
TL;DR: It is concluded that it is the interaction with hepatic lipase rather than the rate of triglyceride transfer that results in the preferred conversion of postprandial A-II-containing HDL2 into HDL3, and that apoA-II exerts a crucial role in this process.
TL;DR: It is demonstrated that Ser153 is involved in the catalytic site of pancreatic lipase and is not crucial for interfacial binding and a Ser-His-Asp catalytic triad similar to that present in serine proteases is present in human pancreaticlipase.
TL;DR: Results indicate that beta-lactoglobulin could participate in the digestion of milk lipids during the neonatal period by enhancing the activity of pregastric lipase through removal of the fatty acids that inhibit this enzyme.
TL;DR: In this article, the Spanish Serrano dry-cured ham was subjected to post-mortem pork muscle and adipose tissue assays and the results showed that the recovered esterase activity was around 50% of the original activity at the end of the drycuring process.
Abstract: Lipase and esterase activities in post-mortem pork muscle and adipose tissue were assayed. Acid lipases showed optimal activity in the presence of 0.8 mg bovine serum albumin (BSA)/ml and 0.05% (by vol.) Triton X-100, while neutral/basic lipases required 5 mg BSA/ml and no addition of Triton X-100. All lipases had an optimal temperature of 37° C except neutral muscle lipase, which was optimally active at 45° C. A wider range of optimal temperatures of 30–45 and 15–45° C was found for muscle acid esterase activity and neutral esterase activity, respectively. In adipose tissue, higher temperatures of 60° C and 45–75° C were found for maximal acid esterase and neutral esterase activities, respectively. Lipolytic and esterolytic activity assays in muscle and adipose tissue were conducted at four different stages in the processing of Spanish Serrano dry-cured ham. Recovered activities of muscle enzymes were more than 40% of the original activity, even at the end of the drycuring process. In adipose tissue, recovered esterase activity was also around 50% of the original activity at the end of the process, while lipolytic activity was significant only during the post-salting stage.