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PubMed1990

Monocyte-specific esterase isoenzyme demonstrated by isoelectric focusing.

Using horizontal thin-layer isoelectric focusing in polyacrylamide gels, we separated the isoenzymes of carboxylic esterase (EC 3.1.1.1) of cell extracts prepared from human hematopoietic cells. Isoenzyme bands were visualized by staining with naphthol ester as substrate and coupling to an azo dye. Staining intensities of isoenzymes were quantified by densitometric scanning. On isoelectric focusing in a pH 2-11 gradient, distinct esterase isoenzyme profiles could be discerned and correlated to various types of normal hematopoietic cells and their leukemic counterparts. One unique isoenzyme, termed monoband, could be clearly identified on the basis of its isoelectric point (pI 6.0), its strong expression by normal and malignant monocytes and its complete and selective inhibition by sodium fluoride. This band was only found in monocytes of either normal or leukemic origin, but not in lymphoid or myeloid cells. The monocyte esterase could be inhibited by sodium fluoride whereas other isoenzyme bands were resistant to this inhibition. However, the specificity of this inhibitory reaction was relative, depending on the concentration of sodium fluoride. Compared with normal monocytes, leukemic monocytes often showed an overexpression of the mono-bands. Dilution experiments established the distinct prominence of the mono-band which could be detected among the other isoenzymes even when only 1% of the total cell population consisted of monocytes. Immature myeloid, but mono-band negative leukemic cells whose arrest of differentiation can be overcome by in vitro 12-O-tetradecanoylphorbol 13-acetate-promoted differentiation to more mature cells, could be induced to express the mono-band which paralleled their maturation to monocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Carboxylesterase↗
PubMedDate not supplied

Identification of arginine esterase as the major androgen-dependent protein secreted by dog prostate and preliminary molecular characterization in seminal plasma.

This work was undertaken to determine the identity of the major androgen-dependent 15,000 molecular weight protein previously observed on SDS polyacrylamide gel electrophoresis of both dog prostate cytosol and dog seminal plasma. The protein was identified as one of the two chains of arginine esterase on the basis of its ability to bind 3H-diisopropylphosphofluoridate (DFP), an active site titrant of serine proteases. Furthermore, since the other polypeptide chain was heterogeneous, at least five distinct peaks of arginine esterase activity could be separated by chromatofocusing under nonreducing conditions. The molecular weight of the seminal plasma protein was estimated at 29,500 by Sephadex G-100 gel filtration, and at 25,000 by SDS polyacrylamide gel electrophoresis in the absence of mercaptoethanol. In the presence of mercaptoethanol, two major peaks were observed with molecular weights of 15,000 and 14,000. These results show that arginine esterase of dog seminal plasma is a serine protease composed of two different chains linked by disulfide bridges. One of the chains contains the reactive serine group. The other one is probably glycosylated since it presents several isoelectric points.

Animals↗
PubMed1999

Human egasyn binds beta-glucuronidase but neither the esterase active site of egasyn nor the C terminus of beta-glucuronidase is involved in their interaction.

Lysosomal beta-glucuronidase shows a dual localization in mouse liver, where a significant fraction is retained in the endoplasmic reticulum (ER) by interaction with an ER-resident carboxyl esterase called egasyn. This interaction of mouse egasyn (mEg) with murine beta-glucuronidase (mGUSB) involves binding of the C-terminal 8 residues of the mGUSB to the carboxylesterase active site of the mEg. We isolated the recombinant human homologue of the mouse egasyn cDNA and found that it too binds human beta-glucuronidase (hGUSB). However, the binding appears not to involve the active site of the human egasyn (hEg) and does not involve the C-terminal 18 amino acids of hGUSB. The full-length cDNA encoding hEg was isolated from a human liver cDNA library using full-length mEg cDNA as a probe. The 1941-bp cDNA differs by only a few bases from two previously reported cDNAs for human liver carboxylesterase, allowing the anti-human carboxylesterase antiserum to be used for immunoprecipitation of human egasyn. The cDNA expressed bis-p-nitrophenyl phosphate (BPNP)-inhibitable esterase activity in COS cells. When expressed in COS cells, it is localized to the ER. The intracellular hEg coimmunoprecipitated with full-length hGUSB and with a truncated hGUSB missing the C-terminal 18-amino-acid residue when extracts of COS cells expressing both proteins were treated with anti-hGUSB antibody. It did not coimmunoprecipitate with mGUSB from extracts of coexpressing COS cells. Unlike mEg, hEg was not released from the hEg-GUSB complex with BPNP. Thus, hEg resembles mEg in that it binds hGUSB. However, it differs from mEg in that (i) it does not appear to use the esterase active site for binding since treatment with BPNP did not release hEg from hGUSB and (ii) it does not use the C terminus of GUSB for binding, since a C-terminal truncated hGUSB (the C-terminal 18 amino acids are removed) bound as well as nontruncated hGUSB. Evidence is presented that an internal segment of 51 amino acids between 228 and 279 residues contributes to binding of hGUSB by hEg.

Amino Acid Sequence↗
PubMed1995

A relatively small 5' regulatory region of esterase S gene of Drosophila virilis determines the specific expression as revealed in transgenic experiments.

Expression of the esterase S gene of Drosophila virilis was studied in transgenic experiments. Truncated genomic copy of this gene including 400 bp of 5' regulatory region was integrated into the genome of Drosophila melanogaster. The products of the transferred gene were detected. It was found that strict temporal and tissue specificity of the esterase S gene expression is conserved in transformed flies. The results suggest that this specificity is evidently determined by the regulatory region of the esterase S gene and controlled by cis mechanism.

Animals↗
PubMed1993

The gut esterase gene (ges-1) from the nematodes Caenorhabditis elegans and Caenorhabditis briggsae.

The ges-1 gene codes for a non-specific carboxylesterase that is normally expressed only in the intestine of the nematode Caenorhabditis elegans. In the current paper, we describe the cloning and characterization of the ges-1 gene from C. elegans, as well as the homologous gene from the nematode Caenorhabditis briggsae. The ges-1 esterases from the two nematodes are 83% identical at the amino acid level and contain regions of significant similarity to insect and mammalian esterases; these conserved regions can be identified with residues believed to be necessary for esterase function. The ges-1 mRNAs from both C. elegans and C. briggsae are trans-spliced. The coding regions, the codon bias and the splicing signals of the two ges-1 genes are quite similar and most (6/7) of the intron positions are retained precisely. Yet, the flanking sequences of the two ges-1 genes appear to have diverged almost completely. For example, the C. elegans ges-1 5'-flanking region (as well as several introns) contains copies of three different SINE-like sequences, previously identified near the hsp-16 genes, near the unc-22 gene and in a repetitive element CeRep-3; none of these elements are found in the C. briggsae ges-1 gene. We show that: (1) the C. elegans ges-1 gene can be used to transform C. briggsae, whereupon expression of the exogenous ges-1 gene is confined to the C. briggsae intestine; (2) the ges-1 homologue cloned from C. briggsae can be transformed into C. elegans, whereupon it is expressed largely in the C. elegans intestine; and (3) a 5'-deletion of the C. elegans ges-1 gene that we have previously shown to be expressed in the C. elegans pharynx is also expressed in the pharynx of C. briggsae (either in the presence or absence of vector sequences). These results suggest that the ges-1 gene control circuits have been maintained between the two nematode species, despite the divergent 5'-flanking sequences of the gene. This raises the question of the evolutionary distance between C. elegans and C. briggsae and we attempt to estimate the C. elegans-C. briggsae divergence time by analysing the rate of synonymous substitutions in coding regions of ges-1 and six other C. elegans-C. briggsae gene pairs. We propose a new method of analysis, which attempts to remove rate differences found between different genes by extrapolating to zero codon bias.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed2001

Three-dimensional structure of Erwinia chrysanthemi pectin methylesterase reveals a novel esterase active site.

Most structures of neutral lipases and esterases have been found to adopt the common alpha/beta hydrolase fold and contain a catalytic Ser-His-Asp triad. Some variation occurs in both the overall protein fold and in the location of the catalytic triad, and in some enzymes the role of the aspartate residue is replaced by a main-chain carbonyl oxygen atom. Here, we report the crystal structure of pectin methylesterase that has neither the common alpha/beta hydrolase fold nor the common catalytic triad. The structure of the Erwinia chrysanthemi enzyme was solved by multiple isomorphous replacement and refined at 2.4 A to a conventional crystallographic R-factor of 17.9 % (R(free) 21.1 %). This is the first structure of a pectin methylesterase and reveals the enzyme to comprise a right-handed parallel beta-helix as seen in the pectinolytic enzymes pectate lyase, pectin lyase, polygalacturonase and rhamnogalacturonase, and unlike the alpha/beta hydrolase fold of rhamnogalacturonan acetylesterase with which it shares esterase activity. Pectin methylesterase has no significant sequence similarity with any protein of known structure. Sequence conservation among the pectin methylesterases has been mapped onto the structure and reveals that the active site comprises two aspartate residues and an arginine residue. These proposed catalytic residues, located on the solvent-accessible surface of the parallel beta-helix and in a cleft formed by external loops, are at a location similar to that of the active site and substrate-binding cleft of pectate lyase. The structure of pectin methylesterase is an example of a new family of esterases.

Amino Acid Sequence↗
PubMed1984

Isoelectric focusing studies of human red cell esterase D: evidence for polymorphic occurrence of a new allele EsD7 in Japanese.

The isoelectric focusing study of esterase D in Japanese revealed evidence of a new polymorphic allele (EsD7) which is difficult to find by conventional starch gel electrophoresis only. A comparison with the occurrence of a subdivision of EsD2 in Caucasians (EsD5) suggests a remarkable difference in allele distribution of esterase D among races. Quantitative analysis showed a relatively low value of enzyme activity for this new allele. It is therefore emphasized that in addition to conventional electrophoresis, enzyme assay and further detection by isoelectric focusing are essential in analyzing the esterase D system.

Alleles↗
PubMed1986

Effect of the esterase-D phenotype on its in vitro enzyme activity.

Esterase-D phenotypes and in vitro activity have been measured in red blood cells from 258 retinoblastoma patients and 73 unaffected relatives. Individuals with the 1-1 and 2-1 phenotypes showed distributions of enzyme activity which were not significantly different from each other. Individuals with the 2-2 phenotype, however, consistently showed a 25-30% lower level of enzyme activity. These results demonstrate the importance of determining the esterase-D phenotype in individuals with low ESD activity who might otherwise be assumed to carry a chromosome deletion at the esterase-D locus. We have also shown that, in vitro, the ESD enzyme is unstable over relatively short periods of time which, if uncontrolled, can give rise to a large variation in measured enzyme levels. The addition of b-mercaptoethanol to the assay buffer, which stabilises the enzyme, results in more consistent values being obtained within the same ESD phenotype. This feature could account in part for much of the variability in enzyme activity observed between different individuals in other studies.

Alleles↗
PubMed1986

Deletions of the esterase D locus from a survey of 200 retinoblastoma patients.

Esterase D levels from 200 retinoblastoma patients have been measured in an attempt to identify individuals carrying deletions of chromosome region 13q14. In this series 75% had bilateral tumours and 23% were familial. Of nine patients identified as having low esterase D levels, five had not previously been diagnosed as deletion carriers. These observations demonstrate the benefit of screening retinoblastoma populations for esterase D deficiency.

Adult↗
PubMed1977

Genetic studies of the Macushi and Wapishana Indians. I. Rare genetic variants and a "private polymorphism' of esterase A.

Blood samples from 509 Macushi and 623 Wapishana Amerindians of of Northern Brazil and Southern Guyana have been analyzed with reference to the occurrence of rare variants and genetic polymorphisms of the following 25 systems: (i) Erythrocyte enzymes: acid phosphatase-1, adenosine deaminase, adenylate kinase-k, carbonic anhydrase-1, carbonic anhydrase-2, esterase A1,2,3, esterase D, galactose-1-phosphate uridyltransferase, isocitrate dehydrogenase, lactate dehydrogenase, malate dehydrogenase, nucleoside phosphorylase, peptidase A, peptidase B, phosphoglucomutase 1, phosphoglucomutase 2, phosphogluconate dehydrogenase, phosphohexoseisomerase, triosephosphate isomerase and (ii) Serum proteins: albumin, ceruloplasmin, haptoglobin, hemoglobin A2 and transferrin. Fifteen different rare variants were detected, involving 11 of these systems. In addition, a previously undescribed variant of ESA 1,2,3 which achieves polymorphic proportions in both these tribes is described. Excluding this variant, the frequency of rare variants is 1.1/1000 in 12510 determinations in the Macushi and 4.7/1000 in 15396 determinations in the Wapishana. The ESA 1,2,3 polymorphism was not observed in 382 Makiritare, 232 Yanomama, 146 Piaroa, 404 Cayapo, 190 Kraho and 112 Moro. Irregularities in the intratribal distribution of this polymorphism in the Macushi and Wapishana render a decision as to the tribe of origin impossible at present. Gene frequencies are also given for previously described polymorphisms of 5 systems: haptoglobin, phosphoglucomutase 1, erythrocyte acid phosphatase, esterase D, and galactose-1-phosphate-uridyl-transferase.

Blood Proteins↗
PubMed1984

Heterogeneity of Es-1 esterases in the rabbit (Oryctolagus cuniculus).

Analysis of the Es-1 system in the rabbit with polyacrylamide gel electrophoresis (PAGE) revealed a high degree of individual variation. In the liver the number of esterase bands found in the Es-1 region of the gels ranged from 2 to 16. The results indicate that one locus with three alleles is responsible for all of the esterase bands in the Es-1 region. The most plausible explanation for the observed heterogeneity is that each of the alleles codes for a protein (MW 65,000 +/- 2000) that is changed by posttranslational modifications, thus giving rise to two to five monomeric enzymes with esterase activity. Polymerization of these monomers then results in 1-11 dimers. Based on similarities with mouse Es-9, chromosomal homology between rabbit Es-1 and mouse Es-9 is proposed.

Animals↗
PubMed1996

Biochemical characterization of juvenile hormone esterases from lines selected for high or low enzyme activity in Gryllus assimilis.

In a previous study, activity of the insect endocrine regulator juvenile hormone esterase (JHE), in the cricket Gryllus assimilis, was subjected to bidirectional selection. This resulted in three pairs of high- and low-selected lines, each of which differed by 3.5-fold in JHE activity. In the present study, juvenile hormone esterases from these lines were characterized with respect to the Michaelis constant (K(m)), thermostability, and inhibition. None of three high-selected JHEs differed from its respective low-selected JHE in the Michaelis constant (K(m)) for juvenile hormone. Similarly, the high-selected JHEs did not differ from the low selected JHEs in thermostability or inhibition by either of two general esterase inhibitors (DFP, eserine) or a "JHE-specific" inhibitor (OTFP). Thus no evidence was obtained to suggest that the response to selection was due to allozymes or isozymes with altered kinetic or stability properties. Kinetic and stability properties were also very similar for the JHEs from the three high-selected or the three low-selected lines. Finally, none of the thermostability or inhibition profiles for any of the six JHEs exhibited sharp discontinuities, thus providing no evidence for the existence of multiple isozymes. The available evidence points to genetically variable regulators which affect the synthesis, degradation, or tissue distribution of JHE as being responsible for the divergence in JHE activity between the selected lines.

Acetone↗
PubMed1991

Genetic characterization of esterase 28 (ES-28) of the house mouse.

The genetics of esterase-28, the major esterase of cauda epididymidis of the house mouse, has been studied after separation by polyacrylamide gel electrophoresis and isoelectric focusing. Four phenotypes are distinguished. Segregation of Es-28 in two backcross series indicated linkage to Es-1, Es-9, and Es-22. The Es-28 locus was placed into esterase cluster 1 on chromosome 8.

Alleles↗
PubMed1986

Effects of a phorbol ester and diacylglycerols on secretion of mucin and arginine esterase by rat submandibular gland cells.

The effects of a phorbol ester, 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and a diacylglyceride, 1-oleoyl-2-acetyl-glycerol (OAG) on the secretion of two major exocrine products by dispersed rat submandibular cells were investigated. TPA stimulated the release of acinar cell mucin and ductal cell protease (arginine esterase) in a dose- and time-dependent manner. Mucin secretion was also provoked by OAG, which, however, had no effect on arginine esterase release. The unsaturated diacylglycerol, 1,2-diolein, elicited a greater mucosecretory response than did OAG at the same concentration, while the saturated 1,2-distearin produced a smaller response. Mucin and enzyme secretion caused by TPA or OAG in the rat submandibular model was not inhibited by either of two putative antagonists, the antipsychotic drug, fluphenazine, and the antibiotic, polymyxin B. The involvement of extracellular Ca2+ in TPA-induced secretion was examined by comparing responses of cells maintained in normal or Ca2+-free medium, or in medium containing the ionophore A23187. Although extracellular Ca2+ was not an absolute requirement for a secretory response, the results indicate a synergistic relationship between TPA and Ca2+ in stimulating the release of both mucin and arginine esterase. These results suggest a role for the Ca2+-, phospholipid-dependent enzyme, protein kinase C in the secretory mechanism of mucous and serous cells in the submandibular gland. This is consistent with the proposal that receptor-mediated hydrolysis of membrane phosphoinositides is an initial event in stimulus-response coupling in exocrine cells.

Animals↗
PubMed1977

The role of acid esterase in the pathogenesis of xanthoma. A histochemical study in rabbits.

Previous studies of our group indicated that two types of cells are involved in the pathogenesis of atheroma. The one type, myocytes, are poor in acid esterase activity, store lipid droplets which they cannot hydrolyze and are transformed into foam cells. The other type, macrophages, are rich in this enzyme activity, can and do hydrolyze the esters with possible healing by fibrosis. In the present study the importance of acid esterase activity in experimental xanthoma in hyperlipidemic rabbits was studied. Perithelial cells of small blood vessels were found to be poor in acid esterase activity and to contain bulky lipid masses. With time, increasing numbers of cells with high enzyme activity were found mainly at a distance from the vessels. These cells, which mostly contained finely emulsified lipids are believed to be macrophages. Thus, in both atheroma and xanthoma, vessel wall cells take a "passive" part in formation of the lesion, while blood borne cells play a role in disposing of the lipid with eventual possibility of healing.

Animals↗
PubMedDate not supplied

Non-invasive sampling for detection of genital infection with Chlamydia trachomatis in males utilising urinary leukocyte esterase tests and immunoassays.

First-void urine specimens from 224 male recruits and 443 patients of venereal disease clinics without complaints of symptoms of urethritis were collected. Urinary leukocyte esterase test, two enzyme immunoassays (EIAs: Syva MicroTrak and Orion), a chemiluminometric assay (Magic Lite) and Syva's MicroTrak direct immunofluorescence test were used. The prevalence of chlamydial urethritis in the study population as determined by direct immunofluorescence test of first-void urine in the military recruits and venereal disease patients was 1.3% and 6.3%, respectively. The denominator used for calculation of sensitivities was the sum of patients with positive test results in at least two of the different test systems used. The sensitivities of first-void urine were 100% for Syva EIA, 96.7% for Orion EIA and 86.7% for the chemiluminometric assay. All assays proved highly specific (99.5-99.7%). Compared with direct immunofluorescence test of first-void urine, the urine leukocyte esterase test had a sensitivity of 93.6% and a specificity of 94.3%. The study showed that the urine leukocyte esterase test is an effective method to detect males infected by Chlamydia trachomatis.

Adolescent↗
PubMed1988

Esterase-16 (ES-16) of the rat (Rattus norvegicus): identification and genetic characterization.

Esterase-16, an esterase present in lung and other tissues of the laboratory rat, has been characterized by its biochemical properties (electrophoretic mobility, substrate pattern, sensitivity to inhibitors) and genetic variation in 107 inbred strains and substrains including 14 RI strains. It was classified as a carboxylesterase (EC 3.1.1.1). The phenotype ES-16A (BN/Han and 63 other strains) was defined as a narrow electrophoretic band migrating between ES-1A and ES-13A, ES-16B (LEW/Han and 42 other strains) exhibited the same electrophoretic mobility as ES-16A but was distinguished by its extremely weak activity. Segregation of ES-16 in RI strains and backcrosses indicated linkage to linkage group V (LGV). The Es-16 locus was tentatively placed into esterase cluster 2 and homology with Es-7 of the house mouse is proposed.

Animals↗