Showing papers on "Esterase published in 1988"

Molecular evidence that insecticide resistance in peach-potato aphids (Myzus persicae Sulz.) results from amplification of an esterase gene.

Abstract: cDNA clones for the esterase (E4) responsible for broad insecticide resistance in peach-potato aphids (Myzus persicae Sulz.) were isolated and used to study the molecular basis of resistance. Increased esterase synthesis by resistant aphids was found to be associated with amplification of the structural gene for the esterase (E4 or its closely related variant, FE4), the degree of amplification being correlated with the activity of the esterase and the level of resistance. Hybridization of the cDNA clones to genomic Southern blots showed that only some of the esterase-related restriction fragments are amplified. Qualitative differences between restriction patterns in different clones of resistant aphids correlated with the presence or absence of a specific chromosome translocation and with production of E4 or FE4.

The E3 protein of bovine coronavirus is a receptor-destroying enzyme with acetylesterase activity.

Abstract: In addition to members of the Orthomyxoviridae and Paramyxoviridae, several coronaviruses have been shown to possess receptor-destroying activities. Purified bovine coronavirus (BCV) preparations have an esterase activity which inactivates O-acetylsialic acid-containing receptors on erythrocytes. Diisopropyl fluorophosphate (DFP) completely inhibits this receptor-destroying activity of BCV, suggesting that the viral enzyme is a serine esterase. Treatment of purified BCV with (/sup 3/H)DFP and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the proteins revealed that the esterase/receptor-destroying activity of BCV is associated with the E3 protein was specifically phosphorylated. This finding suggests that the esterase/receptor-destroying activity of BCV is associated with the E3 protein. Furthermore, treatment of BCV with DFP dramatically reduced its infectivity in a plaque assay. It is assumed that the esterase activity of BCV is required in an early step of virus replication, possible during virus entry or uncoating.

Ferulic Acid Esterase Activity from Schizophyllum commune.

Abstract: Schizophyllum commune produced an esterase which released ferulic acid from starch-free wheat bran and from a soluble ferulic acid-sugar ester that was isolated from wheat bran. The preferred growth substrate for the production of ferulic acid esterase was cellulose. Growth on xylan-containing substrates (oat spelt xylan and starch-free wheat bran) resulted in activity levels that were significantly lower than those observed in cultures grown on cellulose. Similar observations were made for endoglucanase, p-nitrophenyllactopyranosidase, xylanase, and acetyl xylan esterase. Of the enzymes studied, only arabinofuranosidase was produced at maximum levels during growth on xylan-containing materials. Ferulic acid esterase that had been partially purified by DEAE chromatography released significant amounts of ferulic acid from wheat bran only in the presence of a xylanase-rich fraction, indicating that the esterase may not be able to readily attack high-molecular-weight substrates. The esterase acted efficiently, without xylanase addition, on a soluble sugar-ferulic acid substrate.

An acetyl esterase of Trichoderma reesei and its role in the hydrolysis of acetyl xylans

Abstract: An acetyl esterase was purified from Trichoderma reesei by cation and anion exchange chromatography. The enzyme had a molecular weight of 45 000 as determined by SDS-electrophoresis, or 67 000 as determined by gel filtration. In chromatofocusing the enzyme was shown to consist of two isoenzymes with isoelectric points of 6.8 and 6.0. The enzyme showed activity towards naphthyl acetate, triacetin and glucose-and xylose acetates. However, it liberated acetic acid from acetylated xylo-oligomers only to a small extent. The liberation of acetic acid from the oligomeric substrate was enhanced by addition of endoxylanase and β-xylosidase.

Production of acetyl xylan esterase by Trichoderma reesei and Schizophyllum commune

Abstract: Activity of acetyl xylan esterase, an enzyme that removes acetyl groups from acetyl xylan, was coproduced with that of endoxylanase and endoglucanase in two Trichoderma reesei strains and one Schizophyllum commune strain. The levels of activity of extracellular enzymes were measured during the course of cultivation on different carbon sources. The highest activity levels of acetyl xylan esterase were produced by T. reesei QM 9414 in a xylan plus cellulose medium and by S. commune in a cellulose medium. Both strains produced low levels of acetyl xylan esterase activity in glucose, xylose, and cellobiose media. Schizophyllum commune also produced low levels of acetyl xylan esterase activity in xylan and acetyl xylan media. Trichoderma reesei RUT C-30 behaved like a catabolite repression resistant mutant and produced higher enzyme levels than the QM 9414 strain on all carbon sources examined. Analytical gel electrophoresis and isoelectric focusing demonstrated that the acetyl xylan esterase activity of S. co...

Esterase activities in Butyrivibrio fibrisolvens strains.

Abstract: Thirty strains of Butyrivibrio fibrisolvens isolated in diverse geographical locations were examined for esterase activity by using naphthyl esters of acetate, butyrate, caprylate, laurate, and palmitate. All strains possessed some esterase activity, and high levels of activity were observed with strains 49, H17c, S2, AcTF2, and LM8/1B. Esterase activity also was detected in other ruminal bacteria (Bacteroides ruminicola, Selenomonas ruminantium, Ruminobacter amylophilus, and Streptococcus bovis). For all B. fibrisolvens strains tested, naphthyl fatty acid esterase activity paralleled culture growth and was predominantly cell associated. With strains 49, CF4c, and S2, the activity was retained by protoplasts made from whole cells. Esterase activity was detected with all strains when grown on glucose, and some strains showed higher activity levels when grown on other substrates (larchwood xylan or citrus pectin). When nitrophenyl esters of fatty acids were used to measure esterase activity, generally four- to sevenfold-higher activity levels were detected, and with a number of strains substantial levels were found in the culture fluid. Cultures of these strains (H17c, NOR37, D1, and D30g) contained xylanase and acetyl xylan esterase activities, neither of which was associated to any great extent with the cells. Acetyl xylan esterase has not been previously detected in ruminal bacteria and may be important to overall digestion of forage by these organisms.

Purification and characterization of a heat-stable esterase from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius.

Abstract: A heat-stable esterase has been purified 1080-fold to electrophoretic homogeneity from Sulfolobus acidocaldarius, a thermoacidophilic archaebacterium; 20% of the starting activity is recovered. The purified enzyme shows a specific activity of 158 units/mg, based on the hydrolysis of p-nitrophenyl acetate. The esterase hydrolyses short-chain p-nitrophenyl esters, aliphatic esters and triacylglycerols. It is strongly inhibited by paraoxon and phenylmethanesulphonyl fluoride, but only weakly by eserine. From sedimentation-equilibrium data and molecular sieving in polyacrylamide gels, the Mr of the esterase is estimated to be 117000-128000. SDS/polyacrylamide-gel electrophoresis reveals a single band of protein, of Mr 32000. The purified esterase crystallizes in the presence of poly(ethylene glycol) in short rods. The enzyme is inactivated only on prolonged storage at temperature above 90 degrees C.

A highly selective ester hydrolase from Pseudomonas sp. for the enzymatic preparation of enantiomerically pure secondary alcohols; chiral auxiliaries in organic synthesis

Abstract: A series of valuable chiral auxiliaries, (R)- and (S)-(1)–(11), have been prepared in high chemical and optical yields by enzymatic hydrolysis of their esters in the presence of a lipase from Pseudomonas sp.

Purification and partial characterization of poliovirus protease 2A by means of a functional assay.

Abstract: The purification of poliovirus protease 2A from infected cells by a functional assay is described. A small synthetic peptide was cleaved specifically by an esterase present in poliovirus-infected cells. Since the enzyme proved extremely unstable in crude extracts a rapid and efficient purification procedure had to be developed. By treatment with different detergents followed by high-speed centrifugation, the esterase activity was separated from inactivating cellular enzymes and was solubilized. Purification to more than 90% homogeneity could be achieved by a single chromatography step, namely, by gel filtration through Superose 12 under fast-protein liquid chromatography conditions. The esterase activity was associated with a protein of 17,000 daltons and copurified with poliovirus protein 2A. Furthermore, antibodies to 2A specifically precipitated the esterase activity. Thus, the esterase was identified as poliovirus protease 2A. Inhibition studies with known protease inhibitors revealed that 2A is probably a sulfhydryl protease. Of the metal ions tested, only zinc exerted significant inhibitory effects. The esterase activity was optimal near neutral pH and had an extremely short half-life at physiological temperatures.

Preliminary identification of date palm cultivars by esterase isoenzymes and peroxidase activities

Abstract: Two types of crude extracts were obtained from 10 cultivars of date palm (Phoenix dactylifera L.); the Tris buffer extracts from acetone powder were assayed for peroxidase activity and the phosphate buffer extracts were subjected to polyacrylamide gel electrophoresis. The gels were stained for esterase activity using α-naphthylacetate and α-naphthylbutyrate as substrates. The cultivars were divided into four sets on the basis of their esterase isoenzyme phenotypes. These sets were further separated into their individual cultivars on the basis of peroxidase activities. The results are discussed with respect to the possible relationship of the peroxidase activity in the vascular fusariosis of date palm (Bayoud disease). This study has potential for practical application; however, some additional refinements in procedures appear necessary to reduce extraction time.

Two Types of Lipases in Hydrolysis of Polyester

Abstract: Using three kinds of polyesters, polycaprolactone-diol (I), poly(hexamethylene adipate) (II), and a copolyester of I and II, the effects of the molecular weight (Mn) of polyester on their enzymatic hydrolysis were examined. The degrees of hydrolysis of the copolyester by Rhizopus arrhizus and R. delemar lipases were much higher than those of the two homopolymers over a wide range of Mn. On the other hand, the degrees of hydrolysis of the copolyester by Candida cylindracea and hog pancreas lipases and hog liver esterase were between, or near, those of the two homopolyesters. R. delemar lipase could hydrolyze the polyester moiety of polyurethane which hog pancreas lipase could not attack. However, no difference in the hydrolysis products between R. delemar and hog pancreas lipases could be detected. The effects of the melting points of polyesters on their hydrolysis by lipases are discussed.

Molecular cloning and nucleotide sequence of the pectin methyl esterase gene of Erwinia chrysanthemi B374.

Abstract: The gene encoding pectin methyl esterase (pme) has been cloned from Erwinia chrysanthemi B374. Expression of pme in Escherichia coli allowed the enzyme to be characterized. Pectin methyl esterase (PME) was found to have an apparent molecular weight of 36,000 Daltons and an isoelectric point of approximately 9.9. The structural gene was sequenced and consists of a 1098-bp open reading frame encoding a polypeptide of 39,318 Daltons, which includes an amino-terminal signal peptide. The isolation of the Erwinia gene provides a simple method for the production of PME free from depolymerizing pectinases thereby extending its potential uses.

Esterase variation and insecticide resistance in Japanese Aphis gossypii

Abstract: Nine clones of Aphis gossypii Glover (Homoptera: Aphididae) collected from five localities in Japan were tested for esterase activity and resistance to two insecticides. These clones were classified into six types according to esterase pattern and activity detected by electrophoresis. Three clones with very high esterase activity (Type-1) were moderately resistant to malathion and very highly so to pirimicarb. Of six clones with moderate esterase activity one clone (T-4a) was susceptible to malathion but highly resistant to pirimicarb, whereas five clones (T-2, 3a, 3b and 4b) were susceptible to both insecticides. Thus malathion resistance is positively correlated with high esterase activity, whereas pirimicarb resistance is not necessarily so. It is suggested that high esterase activity plays an important role in resistance to these insecticides but that another mechanism is also responsible for resistance to pirimicarb. RESUME Variations de l'activite esterase et resistance aux insecticides chez des Aphis gossypii japonais L'activite esterase et la resistance aux insecticides ont ete examinees chez 9 clones de A. gossypii, recoltes dans 5 sites du Japon. Ces clones ont ete groupes en 6 types suivant le comportement et l'activite de l'esterase mis en evidence par electrophorese. 3 clones ont presente une activite esterase tres elevee (type 1) et des resistances, moyenne pour le malathion et tres elevee pour le pirimicarbe. Parmi les 6 clones ayant une activite esterase moderee, un clone (T-4a) etait sensible au malathion mais tres resistant au pirimicarbe, tandis que les 5 autres clones (T-2, 3a, 3b et 4b) etaient tres sensibles aux deux insecticides. Ainsi, la resistance au malathion presente une forte correlation avec une activite esterase elevee, sans qu'il n'en soit necessairement de meme pour la resistance au pirimicarbe. Une hypothese est emise suivant laquelle l'activite esterase elevee joue un role important dans la resistance a ces deux insecticides, mais qu'un autre mecanisme intervient aussi dans la resistance au pirimicarbe.

Production by Streptomyces viridosporus T7A of an enzyme which cleaves aromatic acids from lignocellulose

Abstract: The lignocellulose-degrading actinomycete Streptomyces viridosporus T7A produced an extracellular esterase when grown in a mineral salts-yeast extract medium Extracellular esterase activity was first detected during the late stationary phase and typically followed the appearance of intracellular activity When the organism was grown in lignocellulose-supplemented medium, esterase activity was not increased, but lignocellulose-esterified p-coumaric acid and vanillic acid were released into the medium Polyacrylamide gels showed that several extracellular esterases differing in substrate specificity were produced Ultrafiltration was used to concentrate the esterase prior to purification Activity was recovered mostly in the molecular weight fraction between 10,000 and 100,000 Concentrated esterase was further purified by DEAE-Sepharose anion-exchange chromatography to a specific activity 1182 times greater than that in the original supernatant There were seven detectable esterase active proteins in the partially purified enzyme solution Three were similar esterases that may be isoenzymes The partially purified esterase had a pH optimum for activity of 90, a temperature optimum of 45 to 50°C, and a Km and Vmax of 0030 mM and 0097 μmol/min per ml, respectively, when p-nitrophenyl butyrate was the substrate The enzyme was unstable above 40°C but retained activity when stored at 4 or −20°C It lost some activity (20%) when lyophilized Substrate specificity assays showed that it hydrolyzed ester linkages of p-nitrophenyl butyrate, α-naphthyl acetate, α-naphthyl butyrate, and lignocellulose Vanillic and p-coumaric acids were identified as products released from lignocellulose The enzyme is thought to be a component of the lignocellulose-degrading enzyme system of S viridosporus

Benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II from Acinetobacter calcoaceticus. Substrate specificities and inhibition studies.

Abstract: The apparent Km and maximum velocity values of benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II from Acinetobacter calcoaceticus were determined for a range of alcohols and aldehydes and the corresponding turnover numbers and specificity constants were calculated. Benzyl alcohol was the most effective alcohol substrate for benzyl alcohol dehydrogenase. Perillyl alcohol was the second most effective substrate, and was the only non-aromatic alcohol oxidized. The other substrates of benzyl alcohol dehydrogenase were all aromatic in nature, with para-substituted derivatives of benzyl alcohol being better substrates than other derivatives. Coniferyl alcohol and cinnamyl alcohol were also substrates. Benzaldehyde was much the most effective substrate for benzaldehyde dehydrogenase II. Benzaldehydes with a single small substituent group in the meta or para position were better substrates than any other benzaldehyde derivatives. Benzaldehyde dehydrogenase II could also oxidize the aliphatic aldehydes hexan-1-al and octan-1-al, although poorly. Benzaldehyde dehydrogenase II was substrate-inhibited by benzaldehyde when the assay concentration exceeded approx. 10 microM. Benzaldehyde dehydrogenase II, but not benzyl alcohol dehydrogenase, exhibited esterase activity with 4-nitrophenyl acetate as substrate. Both benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II were inhibited by the thiol-blocking reagents iodoacetate, iodoacetamide, 4-chloromercuribenzoate and N-ethylmaleimide. Benzyl alcohol or benzaldehyde respectively protected against these inhibitions. NAD+ also gave some protection. Neither benzyl alcohol dehydrogenase nor benzaldehyde dehydrogenase II was inhibited by the metal-ion-chelating agents EDTA, 2,2'-bipyridyl, pyrazole or 2-phenanthroline. Neither enzyme was inhibited by a range of plausible metabolic inhibitors such as mandelate, phenylglyoxylate, benzoate, succinate, acetyl-CoA, ATP or ADP. Benzaldehyde dehydrogenase II was sensitive to inhibition by several aromatic aldehydes; in particular, ortho-substituted benzaldehydes such as 2-bromo-, 2-chloro- and 2-fluoro-benzaldehydes were potent inhibitors of the enzyme.

Characterization of juvenile hormone hydrolysis in early larval development of Trichoplusia ni

Abstract: Extensive juvenile hormone (JH) hydrolysis was detected and characterized in whole-body homogenates of larvae and tissues of Trichoplusia ni during periods of early larval development. The capacity to hydrolyze JH that exists in homogenates of penultimate-instar larvae is far in excess of the measured hormone levels. The major initial metabolites of JH found in diluted homogenates of early-instar larvae and larval tissues were JH acid and JH diol as shown by thin-layer chromatography and microchemical derivatization. Experiments using subcellular fractionation or immunoprecipitation and inhibition studies showed the two hydrolytic activities to be roughly equivalent but located in different subcellular compartments. JH epoxide hydrolase activity was present in the large particle and microsomal fractions, whereas most JH esterase activity was present in the cytosol. Subsequent studies concentrated on JH esterolysis. A titer of JH esterase activity throughout larval development showed this enzyme to be present continuously inside tissues, with periodic manifestations in the hemolymph during each larval molt. Partial purification by affinity chromatography and analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blotting, and isoelectric focusing showed JH esterase from earlyinstar larvae to be indistinguishable from the enzyme from the last instar. Application of JH II or a juvenoid, Ro 10-3108, during any time of early larval development caused no apparent abnormalities, suggesting that the action of JH esterase is not involved with elimination of JH during this period. However, application of a JH esterase inhibitor during a critical period of the third to fourth larval molt caused failure of ecdysis, suggesting that JH acid or at least some esterase or protease may be a factor required for the molt.

Isolation and characterization of sialate 9(4)-O-acetylesterase from influenza C virus.

Abstract: An esterase was isolated from influenza C virus with a specific activity from 1.7-5 U/mg protein, and its substrate specificity was tested with various naturally occurring O-acylated sialic acids, synthetic carbohydrate acetates, and other esters. The enzyme hydrolyses only acetic acid esters at significant rates. The non-natural substrates 4-methyl-umbelliferyl acetate, 4-nitrophenyl acetate, and alpha-naphthyl acetate are cleaved at highest hydrolysis rates, followed by the natural substrate N-acetyl-9-O-acetylneuraminic acid. The esterase also acts on N-glycoloyl-9-O-acetylneuraminic acid and, much slower, on N-acetyl-4-O-acetylneuraminic acid; N-acetyl-7-O-acetylneuraminic acid is not hydrolysed. 2-Deoxy-2,3-didehydro-N-acetyl-9-O-acetylneuraminic acid is also a substrate for this enzyme, however, 6-O-acetylated N-acetylmannosamine and glucose are not. Esterification of the carboxyl function of sialic acids strongly reduces or prevents esterase action on O-acetyl groups. The carboxyl ester is not hydrolysed. The relative cleavage rates also depend on the type of the non-sialic acid part of the molecule. N-Acetyl-9-O-acetylneuraminic acid as component of sialyllactose and rat serum glycoprotein shows hydrolysis rates close to the free form of this sugar, while acetyl ester groups of bovine submandibular gland mucin and rat erythrocytes are hydrolysed at slower rates. Gangliosides and 4-O-acetylated glycoproteins are no substrates for the purified enzyme. A slow hydrolysis is observed by incubation of 9-O-acetylated GD1a with intact influenza C viruses. As other natural acetyl esters (acetyl-CoA and acetylthiocholine iodide) are not hydrolysed, the enzyme can be classified as sialate 9(4)-O-acetylesterase (EC 3.1.1.53).