Development of lipase in nursing piglets

This experiment was designed to investigate the development of gastric lipase and pancreatic lipase in nursing piglets. During the nursing period, the lipase activity was measured at one, 7, 14, 21 and 28 days of age. The results showed that the gastric mucosa weight, pancreas weight, body weight, the specific activity of gastric lipase and pancreatic lipase, and the total activity of gastric lipase and pancreatic lipase increased with the age of the piglets. The development of the specific activity of gastric lipase slowed before the nursing piglets reached 3 weeks of age, but the total activity of gastric lipase at day 28 was significantly higher than that at day 21. The specific activity and total activity of pancreatic lipase were at low levels during the first two weeks of life and then developed quickly from days 21 to 28. It was observed that the specific activity and total activity of gastric lipase were lower than those of pancreatic lipase.     

Evidence for importance of the Staphylococcus hyicus lipase pro-peptide in lipase secretion, stability and activity

Abstract To investigate the function of the pro-peptide (PP) region of the Staphylococcus hyicus exolipase, restriction sites were created in the lipase gene to facilitate the construction of deletions in this region. Lipase gene expression was carried out in Staphylococcus carnosus . In the presence of the entire PP region, the 86-kDa pro-lipase was efficiently exported, had high lipolytic activity, and hardly any degradation products were seen in Western blot analysis. In addition to the 86-kDa pro-lipase, the membrane fraction contained a 106-kDa immunoreactive form. If the PP was completely or partially deleted, signal peptide processing, lipase secretion, lipase activity and/or lipase stability were impaired. The results obtained with lipase PP deletion mutants indicate that the PP region may have two functional domains. The N-terminal region of the lipase PP appears to be more important for lipase activity and the C-terminal portion for lipase secretion and proteolytic stability. In the presence of only the C-terminal part of the PP lipase, secretion was hardly affected. However, the activity of the extracellular lipase was markedly reduced. If only a small portion of the C-terminal part of the PP was present, lipase secretion was again markedly reduced and no lipase activity was detectable. In the presence of the N-terminal half of the PP region, lipase secretion was affected to a lesser extent. However, the resulting 60-kDa form, which showed comparably good specific lipase activity, suffered severe proteolytic degradation.     

Some properties of triacylglycerol lipase in chicken erythrocytes

Membrane-bound acid lipase was found in the chicken erythrocytes ghosts, having an optimum pH of 4.5. The membrane-bound lipase showed its maximum activity at 38 degrees C, and it was stable below 45 degrees C. The bound lipase was activated by octyl glucoside and 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), but it was markedly inhibited by chicken serum. The lipase was solubilized with CHAPS, but the solubilized lipase was labile. The solubilized lipase showed its maximum activity at pH 4.5, 38 degrees C, and it was stable below 40 degrees C. The solubilized lipase was activated by CHAPS and octyl glucoside. The lipase was markedly inhibited by chicken serum. The solubilized lipase have a molecular mass more than 230,000 by Sephacryl S-300 gel filtration.     

Synthesis and secretion of lipoprotein lipase in 3T3-L1 adipocytes. Demonstration of inactive forms of lipase in cells

3T3-L1 adipocytes in culture incorporated [35S]methionine into a protein which could be immunoprecipitated with chicken antiserum to bovine lipoprotein lipase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed this protein had an Mr of 55,000, similar to that of bovine lipoprotein lipase, and accounted for 0.1-0.5% of total protein synthesis in the adipocytes. Lipoprotein lipase protein was present in small amounts in confluent 3T3-L1 fibroblasts, and the amount increased many-fold as the cells differentiated into adipocytes. This increase was accompanied by parallel increases in cellular lipase activity and secretion. When cells were grown with [35S]methionine, the amount of label incorporated into lipoprotein lipase increased for 2 h and then leveled off. Pulse-chase experiments showed that half-life of newly synthesized lipase was about 1 h. Turnover of lipoprotein lipase in control cells involved both release to the medium and intracellular degradation. When N-linked glycosylation was blocked by tunicamycin, the cells synthesized a form of lipase that had a smaller Mr (48,000), was catalytically inactive, and was not released to the medium. Radioimmunoassay demonstrated that 3T3-L1 adipocytes contained an unexpectedly large amount of lipoprotein lipase protein. 55% of the enzyme protein in acetone/ether powder of the cells was insoluble in 50 mM NH3/NH4Cl at pH 8.1, a solution commonly used to extract lipoprotein lipase; 27% of the lipase protein was soluble but did not bind to heparin-Sepharose and had very low lipase activity; and the remaining 13% was soluble, bound to heparin-Sepharose, and had high lipolytic activity. About one-half of the lipase released spontaneously to the medium was inactive, and lipase inactivation proceeded in the medium with little loss of enzyme protein. Lipoprotein lipase released heparin, in contrast, was fully active and more stable. When protein synthesis was blocked by cycloheximide, the level of lipoprotein lipase activity in adipocytes decreased more rapidly than the amount of lipase protein in the cells. Most of the inactive lipoprotein lipase in adipocytes probably results from dissociation of active dimeric lipase, but some could be a precursor of active enzyme.     

Synthesis of inactive nonsecretable high mannose-type lipoprotein lipase by cultured brown adipocytes of combined lipase-deficient cld/cld mice

Combined lipase deficiency (cld) is a recessive mutation which causes a severe deficiency of lipoprotein lipase and hepatic lipase activities and lethal hypertriacylglycerolemia within 3 days in newborn mice. The effect of this genetic defect on lipoprotein lipase was studied in primary cultures of brown adipocytes derived from tissue of newborn mice. Cells cultured from cld/cld mice replicated, accumulated triacylglycerol, and differentiated into adipocytes at normal rates. Lipoprotein lipase activity in unaffected cells was detectable on Day 0 of confluence and increased to 1.3 units/mg DNA by Day 6, while that in cld/cld cells was less than 4% of that in unaffected cells on Days 4-6. Unaffected cells released 1.2% of their lipase activity in 30 min in the absence of heparin, and 11% in 10 min in the presence of heparin, whereas cld/cld cells released no lipase activity. cld/cld cells contained 2-3 times as much lipoprotein lipase protein as unaffected cells, and released no lipase protein to the medium. Immunofluorescent lipoprotein lipase was not detectable in unaffected adipocytes unless lipase secretion was blocked with monesin, causing retention of the lipase in Golgi. cld/cld adipocytes, in contrast, contained immunofluorescent lipoprotein lipase distributed in a diffuse reticular pattern, indicating retention of lipase in endoplasmic reticulum. Lipoprotein lipase immunoprecipitated from cells incubated 1-3 h with [35S]methionine was digested with or without endoglycosidase H (endo H) or F, and resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Lipoprotein lipase in unaffected cells (Mr = 56,000-58,000) consisted of three glycosylated forms, of which the most prevalent was endo H-resistant, the next was totally endo H-sensitive, and the least was partially endo H-sensitive. In contrast, lipoprotein lipase in cld/cld cells (Mr = 56,000) consisted of a single, totally endo H-sensitive form. Lipoprotein lipase in both groups of cells contained two oligosaccharide chains. Chromatography studies with heparin-Sepharose indicated that at least some of the lipoprotein lipase in cld/cld cells was dimerized. The findings demonstrate that brown adipocytes cultured from cld/cld mice synthesize lipoprotein lipase with two high mannose oligosaccharide chains, but it is inactive and retained in endoplasmic reticulum. Whether the cld mutation affects primarily processing of oligosaccharide chains of lipoprotein lipase in endoplasmic reticulum, transport of the lipase from the reticulum, or some other process, is to be resolved.     

介孔分子筛MCM-41固定中性脂肪酶条件的研究

以介孔分子筛MCM-41材料为载体,采用物理吸附法对中性脂肪酶进行了固定化处理,并研究不同条件对固定化脂肪酶催化活性的影响,从而得到该种材料对脂肪酶的最佳固定化条件。给酶量为45960 U/g,固定化温度为45℃,pH值为7.5,时间为3 h,此时固定化酶的活力约为4666 U/g。固定化酶和游离酶的最适反应温度都为40℃,最适pH值为7.5,比游离酶低。固定化酶温度稳定性和pH稳定性较游离酶有所提高。     

Selective release of extrahepatic lipoprotein lipase by polymetaphosphate

We have studied the lipase released into the circulation by polymetaphosphate injection into rats. Lipase release was in proportion to the dose injected. The post-polymetaphosphate lipase was almost completely inhibited by high salt concentrations or by addition of protamine sulfate to the assay system suggesting that this compound released lipoprotein lipase and not hepatic triglyceride lipase. The lipases released by polymetaphosphate and by heparin were compared using a heparin-sepharose affinity column technique which separates lipoprotein lipase from hepatic triglyceride lipase. While heparin released both lipoprotein lipase and hepatic triglyceride lipase, polymetaphosphate released almost exclusively lipoprotein lipase. Other experiments showed that neither polymetaphosphate nor heparin inhibited the hepatic lipase when added to the assay. These results suggest that lipoprotein lipase may be released by the negative charge on these high-charge polymers while hepatic triglyceride lipase release may require the specific sugar configuration of heparin.     

Heparin-releasable lipase activity of rat adrenals, ovaries and testes.

The presence of NaCl-resistant, neutral triacylglycerol hydrolase (lipase) activity in rat adrenal gland, ovary and testis was studied. Both adrenals and ovaries but not testes were found to contain such a lipase. The activity of the enzyme in the adrenal gland was lowered during cortisol treatment and hypothyroidism. An elevated adrenal lipase activity was found during hyperthyroidism. Pseudo-pregnant and lactating rats had higher ovarian lipase activities than cyclic rats. Ovarian lipase activity in lactating rats was positively correlated with the serum concentrations of progesterone and of 20 alpha-hydroxyprogesterone and negatively correlated with the high-density-lipoprotein non-esterified cholesterol concentration. The lipase activity of adrenals and of ovaries was largely releasable from these organs by heparin and could be inhibited by an antibody against heparin-releasable liver lipase. This indicated that the lipase is extracellularly located and is similar to 'liver' lipase. A possible role of this lipase in adrenals and ovaries is discussed.     

Endogenous plasma lipoprotein lipase activity in fed and fasting rats may reflect the functional pool of endothelial lipoprotein lipase

In this study, a correlation was sought between the circulating lipoprotein lipase activity and nutritional state in the rat. In fed rats, the plasma lipoprotein lipase activity was between 30 and 120 munits/ml, whereas after an overnight fast in restraining cages, the lipoprotein lipase plasma levels were between 280 and 500 munits/ml. The plasma lipoprotein lipase activity was inhibited by a specific high titre goat antiserum to rat lipoprotein lipase. No effect of fasting was seen on the plasma hepatic triacylglycerol lipase. 6 h after fasting, adipose tissue lipoprotein lipase decreased maximally, but plasma lipoprotein lipase was not changed and rose only after 16 h. Thus, it seems that most of the lipoprotein lipase activity in the fasting plasma was related to the 3-fold rise in lipoprotein lipase activity in the heart, which may represent total muscle lipoprotein lipase. The increase in heart lipoprotein lipase was due in part to an increase in the t1/2 of the enzyme from 1.2 to 2.9 h. To determine whether the high plasma levels in the fasting rats might result from impaired clearance of the enzyme by the liver, functional hepatectomy was carried out. 15 min after hepatectomy, plasma lipoprotein lipase rose up to 20-fold in fed and about 6-fold in fasting rats. Lipoprotein lipase activity extracted by the liver was calculated to be 30-60 munits/ml in the fed and 171-247 munits/ml plasma per min in fasting rats. An increase in lipoprotein lipase activity in extrahepatic tissues (heart, lung, kidney, diaphragm and adrenal) occurred 30 min after hepatectomy in fed rats. The increase in heart lipoprotein lipase was due to an increase in heparin-releasable fraction. Since no impairment of hepatic clearance of circulating plasma lipoprotein lipase was found, the high fasting plasma lipoprotein lipase activity may be related to an increase in enzyme synthesis, decreased enzyme turnover and an expansion of the functional pool in tissues such as the heart and probably muscle. The present findings indicate that measurement of endogenous plasma lipoprotein lipase can provide information with respect to the size of the functional pool under normal and pathological conditions.     

K55R与ep8叠加突变对扩展青霉脂肪酶热稳定性的改善

利用重叠延伸PCR对扩展青霉脂肪酶(PEL)基因进行体外定点突变,构建了K55R与随机突变体ep8叠加突变的重组质粒pAO815-ep8-K55R。将该质粒电转化引入毕赤酵母(Pichia pastoris)GS115,进行异源表达。实验结果表明:该叠加突变体在毕赤酵母中获得了活性表达,得到表达产物脂肪酶PEL-ep8-K55R-GS。其表达量为508u/mL,分别约为野生型脂肪酶PEL-GS(627u/mL)的81%,随机突变脂肪酶PEL-ep8-GS(924u/mL)的55%;其比活力为2309.1u/mg,与随机突变脂肪酶PEL-ep8-GS和野生型脂肪酶PEL-GS的相仿。叠加突变脂肪酶PEL-ep8-K55R-GS的最适作用温度为37℃,与野生型脂肪酶PEL-GS和随机突变脂肪酶PEL-ep8-GS一致;其Tm值为41.0℃,比野生型脂肪酶PEL-GS提高了2.3℃,比随机突变脂肪酶PEL-ep8-GS提高了0.8℃。表明叠加突变脂肪酶PEL-ep8-K55R-GS的热稳定性有了进一步的提高。     

Glyceride lipases in nerve endings of guinea-pig brain and their stimulation by noradrenaline, 5-hydroxytryptamine and adrenaline

1. Combined guinea-pig cortex and cerebellum was shown to contain triglyceride lipase, diglyceride lipase and monoglyceride lipase, which were assayed by the release of [1-(14)C]palmitate from [1-(14)C]palmitoylglycerol esters. Triglyceride lipase and diglyceride lipase were found in all particulate fractions. 2. With osmotically ruptured synaptosomes the rates of release of palmitate from glyceryl tripalmitate and glyceryl dipalmitate were 7-25mumol/h per g of protein and 0.18-0.69mmol/h per g of protein respectively. The logarithm of the rate of hydrolysis of glyceryl monopalmitate increased linearly with the logarithm of protein concentration. The pH optima of triglyceride lipase and diglyceride lipase were between 7 and 8. The pH optimum for monoglyceride lipase was approx. 8. 3. Triglyceride lipase and diglyceride lipase of osmotically ruptured synaptosomes were stimulated by noradrenaline, 5-hydroxytryptamine and adrenaline. Triglyceride lipase of isolated synaptic membranes was stimulated by 0.01-1mm-noradrenaline. Aging of membranes at 0 degrees C decreased activity, which could still be stimulated by noradrenaline. Diglyceride lipase of isolated membranes was stimulated by 1mum-1mm-noradrenaline. The activity of triglyceride lipase in isolated synaptic vesicles was diminished by 1mm-5-hydroxytryptamine.     

Synthesis and secretion of active lipoprotein lipase in Chinese-hamster ovary (CHO) cells.

Cultured Chinese-hamster ovary cells (CHO cells) were found to produce and secrete a lipase, which was identified as a lipoprotein lipase by the following criteria. Its activity was stimulated by serum and apolipoprotein CII, and was inhibited by high salt concentration. The lipase bound to heparin-agarose and co-eluted with 125I-labelled bovine lipoprotein lipase in a salt gradient. A chicken antiserum to bovine lipoprotein lipase inhibited the activity and precipitated a labelled protein of the same apparent size as bovine lipoprotein lipase from media of CHO cells labelled with [35S]methionine. The lipase activity and secretion were similar in growing cells and in cells that had reached confluency. Hence, lipoprotein lipase appears to be expressed constitutively in CHO cells and is not linked to certain growth conditions, as in pre-adipocyte and macrophage cell lines. At 37 degrees C, but not at 4 degrees C, heparin increased the release of lipase to the medium 2-4-fold. This increased release occurred without depletion of cell-associated lipase activity, suggesting that heparin enhanced release of newly synthesized lipase.     

Digestion in vitro of erythritol esters by rat pancreatic juice enzymes

The mechanism of the digestion of erythritol esters was determined using rat pancreatic juice and purified pancreatic lipase (EC 3.1.1.3). Conditions of hydrolysis were used that would selectively activate or inactivate nonspecific lipase or lipase. It was shown that erythritol tetraoleate was hydrolyzed by nonspecific lipase but not by lipase. The initial digestion product was a triester, predominantly erythritol-1,2,3-trioleate. Thus, nonspecific lipase preferentially hydrolyzed the ester of a primary alcohol. In contrast to the results obtained with the tetraester, lipase could remove a fatty acid from the triester but the resulting erythritol-2,3-dioleate was not hydrolyzed by lipase. The selectivity of this hydrolysis and the inability to hydrolyze the diester are attributed to the known specificity of this enzyme to act only on esters of primary alcohols. Nonspecific lipase completely hydrolyzed erythritol tetraoleate to free erythritol in a stepwise manner. The relative rates of these reactions were tetraester --> triester --> diester --> monoester --> erythritol Because of the specificity of pancreatic lipase and the lack of specificity of nonspecific lipase it is likely that this latter enzyme is the primary agent for the hydrolysis of erythritol esters in the intact animal.     

Cloning,expression and characterization of a lipase gene (<Emphasis Type="Italic">lip3</Emphasis>) from<Emphasis Type="Italic"> Pseudomonas aeruginosa</Emphasis> LST-03

A lipase gene (lip3) was cloned from the Pseudomonas aeruginosa strain LST-03 (which tolerates organic solvents) and expressed in Escherichia coli. The cloned sequence includes an ORF consisting of 945 nucleotides, encoding a protein of 315 amino acids (Lip3 lipase, 34.8 kDa). The predicted Lip3 lipase belongs to the class of serine hydrolases; the catalytic triad consists of the residues Ser-137, Asp-258, and His-286. The gene cloned in the present study does not encode the LST-03 lipase, a previously isolated solvent-stable lipase secreted by P. aeruginosa LST-03, because the N-terminal amino acid sequence of the Lip3 lipase differs from that of the LST-03 lipase. Although the effects of pH on the activity and stability of the Lip3 lipase, and the temperature optimum of the enzyme, were similar to those of the LST-03 lipase, the relative activity of the Lip3 lipase at lower temperatures (0–35°C) was higher than that of the LST-03 lipase. In the absence of organic solvents, the half-life of the Lip3 lipase was similar to that of the LST-03 lipase. However, in the presence of most of the organic solvents tested in this study (the exceptions were ethylene glycol and glycerol), the stability of the Lip3 lipase was lower than that of the LST-03 lipase.Communicated by H. Ikeda     

Synthesis of hepatic lipase in liver and extrahepatic tissues

Immunoprecipitations of hepatic lipase from pulse-labeled rat liver have demonstrated that hepatic lipase is synthesized in two distinct molecular weight forms, HL-I (Mr = 51,000) and HL-II (Mr = 53,000). Both forms are immunologically related to purified hepatic lipase, but not to lipoprotein lipase. HL-I and HL-II are also kinetically related and represent different stages of intracellular processing. Glycosidase experiments suggest that HL-I is the high mannose microsomal form of the mature, sialylated HL-II enzyme. Hepatic lipase activity was detected in liver and adrenal gland but was absent in brain, heart, kidney, testes, small intestine, lung, and spleen. The adrenal and liver lipase activities were inhibited in a similar dose-dependent manner by hepatic lipase antiserum. Immunoblot analysis of partially purified adrenal lipase showed an immunoreactive band co-migrating with HL-II at 53,000 daltons which was absent in a control blot treated with preimmune serum. Adrenal lipase and authentic hepatic lipase yielded similar peptide maps, confirming the presence of the lipase in adrenal gland. However, incorporation of L-[35S]methionine into immunoprecipitable hepatic lipase was not detected in this tissue. In addition, Northern blot analysis showed the presence of hepatic lipase mRNA in liver but not adrenal gland. The presence of hepatic lipase in adrenal gland in the absence of detectable synthesis or messenger suggests that hepatic lipase originates in liver and is transported to this extrahepatic site.     

Mechanisms for turnover of lipoprotein lipase in guinea pig adipocytes

Guinea-pig adipocytes released lipoprotein lipase activity to the medium without depletion of cell-associated lipoprotein lipase activity. Heparin caused immediate release of 20-25% of the lipase activity to the medium, and also enhanced the continued release. After addition of cycloheximide, cell-associated lipoprotein lipase activity decreased rapidly. Release of lipase activity to the medium continued unabated for about 30 min, but there was little release thereafter. The release accounted for only about 25% of the initial lipoprotein lipase activity in the absence and about 50% in the presence of heparin. In pulse-chase experiments with [35S]methionine, labeled lipoprotein lipase appeared in the medium within 40 min, and most of the release occurred during the first h of chase. In a 4-h chase the total (cells + medium) amount of labeled lipase decreased to 34%. Thus, degradation was a main fate of the lipase. Heparin markedly increased the amount of labeled lipase that was released to the medium and decreased the amount that was degraded. Heparin did not change the time-course for the release, and the amount of labeled lipase degraded was proportional to the amount not released to the medium, indicating that the effect of heparin was primarily on release, not on degradation as such. This study demonstrates that adipocytes synthesize lipoprotein lipase in excess of what is being released, and that the excess is rapidly degraded.     

Distribution of lipoprotein lipase and hepatic lipase between plasma and tissues: effect of hypertriglyceridemia

Lipoprotein lipase and hepatic lipase were measured in rat plasma using specific antisera. Mean values for lipoprotein lipase in adult rats were 1.8-3.6 mU/ml, depending on sex and nutritional state. Values for hepatic lipase were about three times higher. Lipoprotein lipase activity in plasma of newborn rats was 2-4-times higher than in adults. In contrast, hepatic lipase activity was lower in newborn than in adult rats. Following functional hepatectomy there was a progressive increase in lipoprotein lipase activity in plasma, indicating that transport of the enzyme from peripheral tissues to the liver normally takes place. Lipoprotein lipase, but not hepatic lipase, increased in plasma after a fat meal. An even more marked increase, up to 30 mU/ml, was seen after intravenous injection of Intralipid. Plasma lipase activity decreased in parallel with clearing of the injected triacylglycerol. 125I-labeled lipoprotein lipase injected intravenously during the hyperlipemia disappeared somewhat slower from the circulation than in fasted rats, but the uptake was still primarily in the liver. Hyperlipemia, or injection of heparin, led to increased lipoprotein lipase activity in the liver. This was seen even when the animals had been pretreated with cycloheximide to inhibit synthesis of new enzyme protein. These results suggest that during hypertriglyceridemia lipoprotein lipase binds to circulating lipoproteins/lipid droplets which results in increased plasma levels of the enzyme and increased transport to the liver.     

Mouse preheparin plasma contains high levels of hepatic lipase with low affinity for heparin

It was recently noted that newborn mice have much higher lipase activity in plasma than rats or humans, and that most of the activity is due to an enzyme related to the hepatic (heparin-releasable) lipase. Here we report that this lipase is present in plasma of adult mice also. In contrast to the high activity of hepatic lipase, the activity of lipoprotein lipase in plasma was low and similar to that in rats. The source of the plasma lipase was probably the liver, since we could not demonstrate hepatic lipase-like activity in any other organ. When human hepatic lipase was injected into mice, it rapidly disappeared from plasma. Most of the injected lipase located in the liver, and could be released back into circulation by injection of heparin. These results indicate that there are binding sites for hepatic lipase in mouse liver, and suggest that mouse hepatic lipase has an affinity for these sites which is lower than usual. It is currently believed that the endothelial acceptors are heparan-sulfate or similar molecules. Mouse hepatic lipase eluted from heparin-Sepharose at lower salt concentration than rat or human hepatic lipase, demonstrating that it has a relatively low affinity for heparin-like polysaccharides.     

Production, properties and application to nonaqueous enzymatic catalysis of lipase from a newly isolated Pseudomonas strain

A potent bacterium for lipase production was isolated from soil and identified as Pseudomonas species. It produced lipase constitutively. A mutant of this strain with a lipase productivity 3.25-fold higher was obtained by treatment with ultraviolet (UV) and nitrosoguanidine (NTG). Its fermentation condition was optimized to a lipase yield of 87.5 U/ml. The lipase had maximum activity at pH 9.0 and 45 degrees C. It was stable at pHs from 7.0 to 11.0 and below 60 degrees C. The effects of metal ions, surfactants and bile salts were also studied. The lipase was 1,3-specific. In organic solvents, the thermal stability of the lipase was significantly enhanced. Its optimum temperature was also slightly increased. The optimum water activity was found between 0.5 and 0.6. The lipase was successfully applied in organic phase to catalyze the glycerolysis of palm oil for monoglyceride (MG) production, and the enantioselective esterification of (R,S)-2-octanol. The enantioselectivity of the lipase could be enhanced substantially by treatment with an amphipathic.     

Reactive properties of the organic solvent-soluble lipase

In a previous report, the organic solvent-soluble lipase was prepared using a synthetic detergent, didodecyl glucosyl glutamate, and it was estimated that 150 +/- 30 molecules of the detergent were attached to one lipase molecule based on gel permeation chromatography and chemical analysis. In this paper, the reactivity of the organic solvent-soluble lipase was compared with that of the native lipase to study the effect of the surrounding detergent on the thermostability and enzymatic reactivity. The activity of the organic solvent-soluble lipase was preserved in the organic solvents up to a temperature of 50 degrees C as in the case of the native lipase in buffer (pH 7.0). The influence of the chain length of fatty acids of the substrate triacylglycerols on the hydrolysis activities was studied. The organic solvent-soluble lipase hydrolyzed triacylglycerols with longer chains more rapidly than the native lipase. The presence of Ca2+ at 0.1 mM stimulated the activity of the native lipase, whereas Ca2+ at a high concentration inhibited it. On the other hand, even at a low concentration, Ca2+ inhibited the activity of the organic solvent-soluble lipase. These results suggest that the detergent attached to the lipase molecule affected the reactive properties.