Mutual effect of invertase and acid phosphatase from the yeast Saccharomyces cerevisiae on their secretion into culture media

The hypothesis that various extracellular enzymes produced by the yeast Saccharomyces cerevisiae exert a mutual influence on their secretion into the culture medium was tested experimentally. The statistically processed results indicate that extracellular invertase affects the secretion of acid phosphatase, and acid phosphatase affects the secretion of invertase. In addition, the secretion of each of these enzymes was shown to be subject to autoregulation.     

Analysis of acid invertase and comparison with acid phosphatase in the ericoid mycorrhizal fungus Hymenoscyphus ericae (Read) Korf and Kernan

Fractions of acid invertase and acid phosphatase of the ericoid mycorrhizal fungus Hymenoscyphus ericae (Read) Korf & Kernan were compared by column chromatography and polyacrylamide gel electrophoresis. Acid invertase levels were measured during the exponential phase after 14 days growth in pure culture. Most acid invertase was wall associated (50%) with 41% forming an extracellular fraction and 9% a soluble, cytoplasmic fraction. The wall-bound fraction was partially solubilized by 1 M NaCl, bulked with the extracellular fraction and separated by gel filtration into two acid invertase activity peaks. These peaks corresponded closely to two acid phosphatase activity peaks measured in the same eluates. Anion exchange chromatography under a continuous salt gradient separated the invertase and phosphatase isoforms from each other. Non-denaturing polyacrylamide gel electrophoresis demonstrated that the more active isoforms of each enzyme have different electrophoretic properties and are high mannose-type glycoproteins with a high affinity for the lectin, concanavalin A. The results are discussed in terms of the functional aspects of the two enzymes and their cytochemical localization.     

Secretion can proceed uncoupled from net plasma membrane expansion in inositol-starved Saccharomyces cerevisiae

Secretion of acid phosphatase and invertase was examined in an inositol-requiring ino1 mutant of the yeast Saccharomyces cerevisiae. Inositol starvation is known to block plasma membrane expansion, presumably due to restricted membrane phospholipid synthesis. If membrane expansion and extracellular protein secretion are accomplished by the same intracellular transport process, one would expect secretion to fail coordinately with cessation of plasma membrane growth in inositol-starved cells. In glucose-grown, inositol-starved cells, plasma membrane expansion and acid phosphatase secretion stopped coordinately, and intracellular acid phosphatase accumulated. In sucrose-grown, inositol-starved cells, plasma membrane growth halted, but secretion of both acid phosphatase and invertase continued until the onset of inositol-less death. Although glucose-grown and sucrose-grown cells differ in their ability to secrete when deprived of inositol, they exhibited the same disturbances in phospholipid synthesis. Phosphatidylinositol synthesis failed, and its precursors phosphatidic acid and CDP-diglyceride accumulated equally in both cultures. Sucrose-grown yeast cells appear to accomplish normal levels of extracellular protein secretion by an inositol-independent mechanism. In glucose-grown yeasts, both plasma membrane expansion and secretion are inositol dependent.     

Discrimination of Mammalian GPI-Anchored Proteins by Hydropathy and Amino Acid Propensities

A 1.7-kb DNA fragment cloned from Zymomonas mobilis genomic DNA complemented the inability to grow on sucrose of a Sue ^? mutant of Z. mobilis that was deficient in the production of both extracellular levansucrase and invertase. Analysis of the nucleotide sequence of the fragment found two open reading frames (ORFs), both of which did not correspond to the structural gene for the levansucrase or the invertase. By subcloning each ORF into two different Suc ^? mutants of Z. mobilis, it has been found that the first ORF (gene zliE) activates the production of the extracellular levansucrase and invertase, and the second ORF (gene zliS) stimulates the secretion of the two enzymes. Gene zliS might contribute to the secretion of proteins having no signal peptide. The expression of zliE and zliS seemed to be under the control of the same promoter.     

Extracellular breakdown of collagen as affected by lysosomal enzymes during the involution of liver cirrhosis

Electron microscopy and electron histochemistry (exposure to acid phosphatase) were used to study the mechanisms of extracellular degradation of collagen in the liver during involution of experimental cirrhosis. The following results were obtained: extracellular secretion of lysosomal enzymes from hepatocytes and connective tissue cells takes place in liver cirrhosis and its involution; partial hepatectomy during liver cirrhosis stimulates the activity of acid phosphatase in the liver cells; the lysosomal enzymes, excreted from hepatocytes and connective tissue cells by means of exocytosis take an active part in collagen extracellular degradation in vivo; at initial stages of cirrhosis involution extracellular degradation of collagen in the liver occurs at the expense of lysosomal enzymes from hepatocytes and connective tissue cells. Subsequently, as cirrhosis regresses, the principal role in the lysis of collagen gradually passes to lysosomal enzymes of hepatocytes.     

酸雨、铜和莠去津对土壤水解酶活性的影响

运用正交试验设计方案,考察了酸雨、重金属铜和农药莠去津复合污染情况下各因素的主效应及两两交互作用对土壤脲酶、转化酶和酸性磷酸酶活性的影响。结果表明,H^+、Cu^2+对酶活性影响显著且其对3种酶的抑制效应为H^+>Cu^2+;Al^3+仅对脲酶表现出一定的激活效应;而除磷酸酶外,莠去津对脲酶、转化酶活性无明显影响。交互作用试验表明：莠去津×Cu的协同作用主要表现为对磷酸酶活性的影响;而H×Cu对脲酶、转化酶均表现为颉抗作用;在本文所考察的浓度范围内,未见莠去津与氢有显著的交互作用存在3种酶中,以磷酸酶对外来污染物最为敏感。     

Acid phosphatase localization in the fungus Whetzelinia sclerotiorum

Acid phosphatase was localized by light and electron microscopy in chains of vacuoles in hyphal tip cells of Whetzelinia sclerotiorum. The Enzyme was present in these vacuoles whether or not conditions favored extracellular acid phosphatase secretion. Apical vesicles, microbodies, woronin bodies, and lipid bodies did not contain acid phosphatase. The implications regarding terminology of organelles in filamentous fungi are discussed with special reference to the fungal spherosome concept.Abbreviations AP acid phosphatase     

Secretion and localization of invertase and inulinase in recombinant Saccharomyces cerevisiae

Summary Secretion of invertase and inulinase produced by recombinant Saccharomyces cerevisiae cells were investigated under derepression conditions of GALI promoter. Secreted invertase mainly localized in the periplasmic space, but most of inulinase was found in the extracellular culture medium. This high level of extracellular secretion of inulinase was not dependent on the growth phase in which derepression of GALI promoter occurs. Our results indicate that the inulinase polypeptide itself may have a function for the protein secretion into the culture medium.     

Wall-bound enzymes in callus of Convolvulus arvensis

Isolated cell walls of Convolvulus callus contain α- and β-galactosidase, α- and β-glucosidase, α- and β-mannosidase, acid invertase and acid phosphatase activities. No neutral invertase or alkaline phosphatase activities could be detected. Acid invertase activity per mg cell wall increased considerably during incubation of callus fragments in nutrient solution, as opposed to the activities of the other enzymes mentioned.     

Secretory expression of human growth hormone inSaccharomyces cerevisiae using three different leader sequences

A recombinant human growth hormone (hGH) was expressed as a secretory product in the yeastSaccharomyces cerevisiae. Three different leader sequences derived from the mating factor α1 (MFα1), inulinase and invertase were used to direct the secretion of hGH into the extracellular medium. Among three leader sequences tested, the inulinase leader sequence was found to be the most efficient in the secretory expression of hGH. In contrast, no hGH was detected in the extracellular medium with the invertase leader sequence. After 48 h shake-flask culture, the yields of hGH secreted into the medium by the invertase, MFα1, inulinase and invertase leader sequences were approximately 0, 0.3 and 0.9 mg/L, respectively. The secretion efficiencies were also found to be 0, 3.8 and 13% for the invertase, MFα1 and inulinase leader sequences, respectively.     

Acid phosphatase and invertase activities of <Emphasis Type="Italic">Aspergillus niger</Emphasis>

Aspergillus niger is widely used as an enzyme source in industries. Considering its enzymic potential, A. niger was studied for its acid phosphatase (EC 3.1.3.2, orthophosphoric monoester phosphohydrolase), and invertase (EC 3.2.1.26, β-fructofuranoside fructohydrolase) activity in defined media supplemented with 1%, 3%, or 5% sucrose concentrations. Both these enzymes play a key role in phosphate and carbon metabolism in plants, animals, and microorganisms and hence are interesting from the standpoint of biotechnological applications. Ontogenic changes in extracellular, cytoplasmic, and wall-bound enzyme activities of A. niger were studied. Growth in terms of fresh weight showed inverse correlation with pH. At higher pH values, both enzyme activities were higher in the medium supplemented with low sucrose concentration. It was observed that the more the fresh weight of fungi decreased, the greater was the enzyme activity observed. It is suggested that these enzymes may participate in autolysis of fungi and, on the other hand, could prove to be a potential source of industrial application and exploitation.     

An electron microscope study of acid hydrolase activity in the pneumonocytes of Xenopus laevis

Synopsis Electron microscope techniques were used to study the intercellular distribution of aryl sulphatase and acid phosphatase in the pneumonocytes which line the air spaces in the lung ofXenopus laevis. Strong reactions for aryl sulphatase and acid phosphatase were present in the limiting membranes of the cytoplasmic inclusion bodies; no significant activity was found in the inclusion contents or in the membranous material present in the air spaces. Both enzymes were present in the multivesicular bodies but other organelles were unreactive. These results suggest that hydrolytic enzymes are involved in the secretion of surface-active materials in amphibian lung.     

Effect on lysosomes of invertase endocytosed by rat-liver

The intracellular localization of invertase endocytosed by rat liver was investigated by analytical centrifugation in sucrose and Percoll gradients of mitochondrial fractions originating from rats killed 15 h after injection. After isopycnic centrifugation in a sucrose gradient, invertase is located in higher density zones than acid hydrolases. The difference between the distribution of invertase and that of acid hydrolases increases with the amount of invertase injected. When the invertase dose is sufficiently high, a change of lysosomal enzyme distribution is clearly visible. It consists in the shift of a proportion of these enzymes to higher density regions where invertase is located. The proportion of hydrolase activity affected by invertase is different for each enzyme measured; it is the least pronounced for acid phosphatase, and most for acid deoxyribonuclease and arylsulfatase. A pretreatment of the rat with Triton WR 1339 considerably decreases the equilibrium density of structures bearing invertase. Nevertheless invertase distribution is quite distinct from that of the bulk of lysosomal enzymes that are recovered in lower density zones of the gradient; on the other hand the invertase injection to rats treated with Triton WR 1339 causes a spreading of the acid hydrolase distribution towards higher density zones. The distribution of acid hydrolases and invertase in a Percoll gradient depends on the sucrose concentration of the solvent. It is shifted towards higher densities when the sucrose concentration increases. The phenomenon is more important for invertase. These results are best explained by supposing that invertase accumulates in a distinct population of lysosomes that can be individualized as a result of the density increase they are subjected to by the invertase they accumulate. It is proposed that these lysosomes mainly originate from non-parenchymal cells of the liver.     

Acid and alkaline invertases in roots and nodules of Lupinus angustifolius infected with Rhizobium lupini

Summary Both acid and alkaline invertase activity were found in tips and cortical tissue of Lupinus angustifolius L. roots infected with Rhizobium lupini NZP 2257. Only the alkaline invertase was detected in the nodule cytoplasm. Weak invertase activity found in the bacteroids was probably a contamination from plant invertase. The alkaline invertase activity in the nodule cytoplasm was 250 times that detected in the bacteroids and 8 times that detected in cortical tissue. No intracellular or extracellular invertase was detected in R. lupini cultured in liquid medium containing sucrose.     

Differences in invertase activity in embryogenic and non-embryogenic calli from Medicago arborea

The different invertase activities in embryogenic and non-embryogenic calli induced from explants (cotyledons, petioles, hypocotyls and leaves) obtained from Medicago arborea L. subsp. arborea seedlings were evaluated. Total invertase activity was lower in the calli with the greatest embryogenic capacity. The greatest fraction of this activity corresponded to soluble invertase. Wall-bound invertase showed maximum activity during the first two months of culture and the highest activities of this type were found in non-embryogenic calli. Extracellular invertase formed the smallest fraction of the total invertase activity evaluated. Acid and alkaline invertase activities were found in all calli but differences were detected between the embryogenic and non-embryogenic calli. In the former, the activity of both types of invertase exhibited a similar type of behaviour but different from that observed in the non-embryogenic calli. The calli with the greatest embryogenic capacity had very low levels of acid invertase and very high levels of the alkaline form. Soluble invertase – both acid and alkaline – accounted for the highest fraction after the first two months of culture and was present in lower amounts in the embryogenic than in the non-embryogenic calli. Regarding bound invertase, the highest production was seen to correspond to acid invertase. The extracellular invertase evaluated corresponded to the acid form since the alkaline extracellular invertase did not show any physiologically significant activity.     

马尾松人工林土壤有机层和矿质土壤层酶活性随雨旱季的变化

土壤酶活性随雨旱季的变化特征对于深入理解人工林土壤物质循环具有重要意义。2014年6月至2017年4月,采用原位土柱培养方法,研究了长江上游低山丘陵区的马尾松人工林土壤有机层和矿质土壤层转化酶、脲酶和酸性磷酸酶活性随关键时期(雨季初期;雨季中期;雨季末期;旱季初期;旱季末期)的变化特征。结果表明：土壤有机层和矿质土壤层酶活性随关键时期的变化显著,总体表现为雨季高于旱季。土壤有机层转化酶和脲酶活性呈现逐年降低的趋势;矿质土壤层酸性磷酸酶活性呈现逐年增加的趋势。土壤有机层转化酶和酸性磷酸酶活性显著高于矿质土壤层。土壤有机层三种酶活性年际变化幅度及随关键时期的变化幅度均高于矿质土壤层。偏最小二乘分析(PLS)表明,土壤含水量、微生物量和底物含量对土壤酶活性具有显著影响,但其影响大小强烈取决于土壤层次和酶的种类。可见,马尾松人工林土壤有机层酶活性对环境因子变化的响应更敏感,而且驱动土壤有机层和矿质土壤层酶活性动态的主导因子也存在差异。     

Studies on the physiology of hyacinth bulbs (Hyacinthus orientalis L.) the effect of plant growth regulators on metabolic activities in non-chilled hyacinth bulbs

The effect of gibberellic acid, benzyladenine and a mixture of these compounds on the activity of some hydrolytic enzymes was studied in hyacinth bulbs non-exposed to low temperatures. Plant growth regulators were applied on the heel of dormant bulbs in the middle of July. An intensive elongation of the inflorescence was observed only in the plants treated previously with GA₃ and grown in a warm greenhouse for 63 and 84 days. The activities of amylase, invertase and acid phosphatase were higher in the flower buds of unrooted bulbs treated with growth regulators than in the control plants, although growth regulators did not affect the level of extractable proteins. The elongation of the inflorescence in the plants treated with gibberellic acid was correlated with a sharp increase of invertase activity in this organ. The effect of GA₃ and BA on the activity of other enzymes in buds and scales varied with the period of plant growth.     

Processing, transport, and secretion of the lysosomal enzyme acid phosphatase in Dictyostelium discoideum

To explain the different secretion kinetics of lysosomal enzymes in Dictyostelium discoideum, previous investigators have hypothesized the existence of a heterogeneous population of lysosomes containing either the enzyme acid phosphatase or other hydrolase enzymes. This proposal predicts that at least two targeting mechanisms exist for lysosomal enzymes in this organism. To begin to investigate this possibility, the transport, processing, and targeting of acid phosphatase was studied by using a combination of radiolabel pulse-chase procedures, subcellular fractionations, and indirect immunofluorescence microscopy. Acid phosphatase was initially synthesized in axenically growing cells as a 56-kDa precursor polypeptide that was proteolytically processed after 20 min to a 55-kDa mature protein. This enzyme was rapidly transported from the endoplasmic reticulum to Golgi complex (halftime of 3 min) as measured by the acquisition of resistance to the enzyme endoglycosidase H. Furthermore, Percoll gradient fractionations indicated that radiolabeled forms of acid phosphatase reached dense lysosomal vesicles at about the same time as final processing was occurring. Proper sorting of acid phosphatase in D. discoideum apparently was not critically dependent on low intravacuolar pH since the addition of ammonium chloride did not stimulate the missorting and secretion of acid phosphatase. These results are very similar to previous observations concerning other Dictyostelium lysosomal enzymes. Consistent with the existence of a heterogeneus population of lysosomes, the percentage of radiolabeled acid phosphatase secreted 4 h into a chase period was 15-fold lower as compared with another lysosomal enzyme, beta-glucosidase. However, acid phosphatase, alpha-mannosidase, and beta-glucosidase were all predominantly colocalized as determined by indirect immunofluorescence, which for the first time demonstrates the homogeneous nature of the lysosomal system in D. discoideum. Taken together these results suggest that the processing and transport of acid phosphatase may be similar in nature to the glycosidases. However, the different kinetics of secretion of acid phosphatase versus the colocalized glycosidase enzymes suggests that an undefined mechanism operates to distinguish these classes of enzymes at a step after localization to lysosomes but prior to secretion.