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1.
Y Dasarathy J Stevens B L Fanburg J J Lanzillo 《Archives of biochemistry and biophysics》1989,270(1):255-266
A peptidyl dipeptidase-4 (bacterial PDP-4) was purified to near homogeneity from a supernatant of Pseudomonas maltophilia extracellular medium. Bacterial PDP-4 is a single-polypeptide-chain enzyme, 82 kDa, with an alkaline isoelectric point. Peptides susceptible to hydrolysis by bacterial PDP-4 include angiotensin 1, bradykinin, enkephalins, atriopeptin 2, and smaller synthetic peptides. N-acylated tripeptides are hydrolyzed, but free tripeptides are not. A free carboxy terminus is required for hydrolysis. Peptides with ultimate and penultimate Pro residues are not hydrolyzed. The enzyme does not require an anion for activity. Bacterial PDP-4 was inhibited by EDTA and the dipeptide Phe-Arg. Thiorphan was an inhibitor only at levels well above those required for inhibition of neutral metalloendopeptidase (NEP), an enzyme for which thiorphan is specific. A second NEP and thermolysin inhibitor, phosphoramidon, did not inhibit bacterial PDP-4. The potent angiotensin-converting enzyme inhibitor lisinopril was not inhibitory. Bacterial PDP-4 is distinguished from a similar enzyme from Escherichia coli, which is not susceptible to EDTA inhibition, and one from Corynebacterium equi, which hydrolyzes free tripeptides. These data indicate that the bacterial PDP-4 catalytic site is unlike those of other enzymes that function either wholly or in part as peptidyl dipeptidases. 相似文献
2.
Anthony J. Turner John Hryszko Nigel M. Hooper Michael J. Dowdall 《Journal of neurochemistry》1987,48(3):910-916
The electric organ of Torpedo marmorata contains a membrane-bound, captopril-sensitive metallopeptidase that resembles mammalian angiotensin converting enzyme (peptidyl dipeptidase A; EC 3.4.15.1). The Torpedo enzyme has now been purified to apparent homogeneity from electric organ by a procedure involving affinity chromatography using the selective inhibitor lisinopril immobilised to Sepharose via a 28-A spacer arm. The purified protein, like the mammalian enzyme, acted as a peptidyl dipeptidase in cleaving dipeptides from the C-terminus of a variety of peptide substrates, including angiotensin I, bradykinin, [Met5]enkephalin, [Leu5]enkephalin, and the model substrate hippuryl (benzoylglycyl; BzGly)-His-Leu. The hydrolysis of BzGly-His-Leu was activated by Cl-. Enzyme activity was inhibited by classical angiotensin converting enzyme inhibitors, including captopril, enalaprilat (MK422), and lisinopril (MK521). Torpedo angiotensin converting enzyme, like its mammalian counterpart, was also able to act as an endopeptidase in hydrolysing the amidated neuropeptide substance P. Hydrolysis of substance P occurred primarily at the Phe8-Gly9 bond with release of the C-terminal tripeptide, Gly-Leu-MetNH2, and this hydrolysis was blocked by selective inhibitors. The Torpedo enzyme was recognised by a polyclonal antibody to pig kidney angiotensin converting enzyme on immunoelectrophoretic (Western) blot analysis. Thus, on the basis of substrate specificity, inhibitor sensitivity, and immunological criteria, the Torpedo enzyme closely resembles mammalian angiotensin converting enzyme. However, the Torpedo enzyme appears somewhat larger (Mr = 190,000) than the pig kidney enzyme (Mr = 180,000) on sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The endogenous peptide substrate(s) for Torpedo electric organ angiotensin converting enzyme and the physiological role of the enzyme in this tissue remain to be evaluated. 相似文献
3.
Conversion of angiotensin-1 to angiotensin-2 by a latent endothelial cell peptidyl dipeptidase that is not angiotensin-converting enzyme 总被引:1,自引:0,他引:1
J J Lanzillo Y Dasarathy J Stevens B L Fanburg 《Biochemical and biophysical research communications》1986,134(2):770-776
Cultured bovine pulmonary artery endothelial cells contain a second peptidyl dipeptidase, distinct from angiotensin-converting enzyme, present in an inactive form associated with a non-dialyzable inhibitor. Partial purification by glycine affinity chromatography separates enzyme from inhibitor to yield a preparation which hydrolyzes angiotensin-1, bradykinin, substance P, atriopeptin-2, enkephalin and Hip-His-Leu. This enzyme is resistant to inhibition by lisinopril, captopril, thiorphan, phosphoramidon, soybean trypsin inhibitor, PMSF and aminopeptidase and carboxypeptidase inhibitors, but is inhibited by EDTA. 相似文献
4.
The activity of peptidyl dipeptidase (peptidyldipeptide hydrolase, EC 3.4.15.1), also known as angiotensin-converting enzyme, was studied in small blood vessel preparations isolated from rabbit brain. The vascular preparation contained arterioles and capillaries and was essentially free of extravascular material. Enzymatic activity was demonstrated in microvessel homogenates using both hippuryl-histidyl-leucine and tritium-labeled angiotensin I as substrates. Activity in the microvessels was dependent on the presence of chloride ion and was sensitive to inhibition by converting enzyme inhibitors previously shown to be effective in both vivo and in vitro. Specific activity in the micro-vessels was approximately 20 times that in homogenates of brain, and was almost 60% of that found in rat lung homogenates. The data were consistent with an endothelial localication for peptidyl dipeptidase in the cerebral vasculature and supports the proposal that this enzyme has a physiological role in extrapulmonary vascular beds. 相似文献
5.
V L von Riesen 《Applied microbiology》1980,39(1):92-96
Pseudomonas maltophilia and Pseudomonas putida were identified as alginolytic species. Two media used for demonstrating alginolytic activity are described. The applied aspects of the ability of these two species to digest algin are discussed. 相似文献
6.
M P Filatova N A Krit N V Beschastnaia A V Blokhina N I Kozlova 《Bioorganicheskaia khimiia》1985,11(1):21-30
The synthesis of 5 analogues of the effective inhibitor of peptidyl dipeptidase, teprotide, has been carried out. The inhibitory and bradykinin-potentiating activity of these compounds has been assayed. N-Terminal pyroglutamic acid and positive charge of arginine in position 4 were found to be essential for biological activity of the inhibitor. 相似文献
7.
Bacteria belonging to the Pseudomonas genus and isolated from zonal soils in different geographical zones of the USSR as well as from the rhizosphere of cultivated and wild plants were tested for their esterase activity. The studied collection of cultures included 205 strains of different pigmented Pseudomonas species which, according to the conventional taxonomy, were assigned to the so-called "Pseudomonas fluorescens complex". As was shown in this study, many Pseudomonas species are potential producers of nonspecific esterases. P. maltophilia and P. geniculata synthesizing pyomelanin have the highest activity of esterase. The activity of esterase correlates with the formation of a melanin-like pigment in Pseudomonas cultures. It also correlates with the species to which a culture belongs, which makes it possible to use this property as an additional criterion for the identification of Pseudomonas species. 相似文献
8.
Peptidyl dipeptidase activity distinct from the angiotensin converting enzyme (EC 3.4.15.1) was isolated from membrane fractions of rabbit kidney and lung. The enzyme cleaved Leu-enkephalin at the Gly-Phe bond, releasing Tyr-Gly-Gly and Phe-Leu, and also acted on bradykinin releasing the terminal dipeptide Phe-Arg. In contrast to the converting enzyme, however, this peptidyl dipeptidase did not act on angiotensin I, or on hippuryl His-Leu, nor was it inhibited by captopril (SQ 14225) or by SQ 20881. Kinetic studies indicated a Km for the kidney enzyme of 80 μM with Leu-enkephalin as a substrate. Our findings indicate that more than one enzyme is present in membrane preparations of lung and kidney inactivating enkephalin, and suggest a role for these enzymes in the peripheral actions of opiate and related peptides. 相似文献
9.
Membrane peptidases in the pig choroid plexus and on other cell surfaces in contact with the cerebrospinal fluid. 总被引:5,自引:2,他引:3
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A comprehensive survey of 11 peptidases, all of which are markers for renal microvillar membranes, has been made in membrane fractions prepared from pig choroid plexus. Two fractionation schemes were explored, both depending on a MgCl2-precipitation step, the preferred one having advantages in speed and yield of the activities. The specific activities of the peptidases in the choroid-plexus membranes were, with the exception of carboxypeptidase M, lower than in renal microvillar membranes: those of aminopeptidase N, peptidyl dipeptidase A ('angiotensin-converting enzyme') and gamma-glutamyltransferase were 3-5-fold lower, those of aminopeptidase A and endopeptidase-24.11 were 12-15 fold lower, and those of dipeptidyl peptidase IV and aminopeptidase W were 50-70-fold lower. Carboxypeptidase M had a similar activity in both membranes. Alkaline phosphatase and (Na+ + K+)-activated ATPase were more active in the choroid-plexus membranes. No activity for microsomal dipeptidase, aminopeptidase P and carboxypeptidase P could be detected. Six of the peptidases and (Na+ + K+)-activated ATPase were also studied by immunoperoxidase histochemistry at light- and electron-microscopic levels. Endopeptidase-24.11 and (Na+ + K+)-activated ATPase were uniquely located on the brush border, and the other two peptidases appeared to be much more abundant on the endothelial lining of microvessels. Dipeptidyl peptidase IV and aminopeptidase W were also detected in microvasculature. Pial membranes associated with the brain and spinal cord also stained positively for endopeptidase-24.11, aminopeptidase N and peptidyl dipeptidase A. The immunohistochemical studies indicated the subcellular fractionation did not discriminate between membranes derived from epithelial cells (i.e. microvilli) and those from endothelial cells. The possible significance of these studies in relation to neuropeptide metabolism and the control of cerebrospinal fluid production is discussed. 相似文献
10.
Bacterial Antagonists to Verticillium dahliae Kleb. 总被引:7,自引:0,他引:7
Bacteria were isolated from the rhizosphere of Verticillium dahliae hosts and from environments. A total of 1394 bacterial isolates were screened for their ability to inhibit in vitro the growth of the phytopathogenic fungi V. dahliae ; 15% (203 of the isolates) showed antifungal effects. Seventeen isolates were selected and determined for further investigations, seven different species were identified. Several of the bacterial species listed, e.g. Erwinia herbicola, Pseudomonas chlororaphis, Pseudomonas paucimobilis and Xanthomonas maltophilia have not been reported previously as bacterial antagonists of V. dahliae. Bacillus subtilis, Pseudomonas fluorescens and Xanthomonas maltophilia are strong antagonists. We proved that lytic enzymes and siderophores are involved in the inhibition of growth. Ultrastructural and morphological changes were induced in Verticillium dahliae by the antagonistic bacteria. 相似文献
11.
Nucleic Acid Enzyme Studies of Nonfermentative Gram-Negative Bacteria Using Thin-Layer Chromatography 总被引:1,自引:1,他引:0
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A rapid and easy to perform procedure for determining the nucleic acid enzyme reactions of intact bacterial cells was developed. Overnight organism growth on triple sugar agar was tested for nucleoside phosphotransferase, nucleosidase, and nucleotidase activity. Reaction products were detected by means of thin-layer chromatography and fluorescence. Characteristic patterns were seen with certain strains of nonfermentative gram-negative bacteria, which indicated that these tests could aid in classification and identification of E0-1 group, Pseudomonas multivorans, P. cepacia, P. maltophilia, certain other Pseudomonas species, and Herellea vaginicola. 相似文献
12.
目的了解绍兴市人民医院2010年1月到2011年6月嗜麦芽窄食单胞菌的临床分布情况及耐药性分析,为临床合理用药提供依据。方法从近年来该院各感染标本中分离出嗜麦芽窄食单胞菌,用法国梅里埃全自动微生物鉴定仪对细菌进行鉴定,并做临床常用抗生素耐药性分析,用WHONET 5.4软件进行统计学分析。结果 113株分离的嗜麦芽窄食单胞菌主要来自于ICU,标本主要来源于呼吸道感染的痰标本中;药敏结果显示,复方新诺明、米诺环素和左旋氧氟沙星等对嗜麦芽窄食单胞菌有较好的抗菌活性,敏感率依次为96.46%、96.46%和92.92%;对亚胺培南100%耐药;对氨苄西林/舒巴坦、美洛培南和氨曲南的耐药率依次为91.07%、90.27%和89.38%。结论该院临床分离的嗜麦芽窄食单胞菌主要来源于呼吸道感染患者的痰液标本中,以ICU患者感染居多,耐药现象比较严重,应加强耐药性监测,以指导临床合理用药,控制院内感染。 相似文献
13.
The hydrolysis of alpha-human atrial natriuretic peptide by pig kidney microvillar membranes is initiated by endopeptidase-24.11. 总被引:10,自引:4,他引:10
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alpha-Human atrial natriuretic peptide, a 28-amino-acid-residue peptide, was rapidly hydrolysed by pig kidney microvillar membranes in vitro, with a t1/2 of 8 min, comparable with the rate observed with angiotensins II and III. The products of hydrolysis were analysed by h.p.l.c., the pattern obtained with membranes being similar to that with purified endopeptidase-24.11 (EC 3.4.24.11). No hydrolysis by peptidyl dipeptidase A (angiotensin I converting enzyme, EC 3.4.15.1) was observed. The contribution of the various microvillar membrane peptidases was assessed by including specific inhibitors. Phosphoramidon, an inhibitor of endopeptidase-24.11, caused 80-100% suppression of the products. Captopril and amastatin (inhibitors of peptidyl dipeptidase A and aminopeptidases respectively) had no significant effect. Hydrolysis at an undefined site within the disulphide-linked ring occurred rapidly, followed by hydrolysis at other sites, including the Ser25--Phe26 bond. 相似文献
14.
A periplasmic D-alanyl-D-alanine dipeptidase in the gram-negative bacterium Salmonella enterica
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The VanX protein is a D-alanyl-D-alanine (D-Ala-D-Ala) dipeptidase essential for resistance to the glycopeptide antibiotic vancomycin. While this enzymatic activity has been typically associated with vancomycin- and teicoplainin-resistant enterococci, we now report the identification of a D-Ala-D-Ala dipeptidase in the gram-negative species Salmonella enterica. The Salmonella enzyme is only 36% identical to VanX but exhibits a similar substrate specificity: it hydrolyzes D-Ala-D-Ala, DL-Ala-DL-Phe, and D-Ala-Gly but not the tripeptides D-Ala-D-Ala-D-Ala and DL-Ala-DL-Lys-Gly or the dipeptides L-Ala-L-Ala, N-acetyl-D-Ala-D-Ala, and L-Leu-Pro. The Salmonella dipeptidase gene, designated pcgL, appears to have been acquired by horizontal gene transfer because pcgL-hybridizing sequences were not detected in related bacterial species and the G+C content of the pcgL-containing region (41%) is much lower than the overall G+C content of the Salmonella chromosome (52%). In contrast to wild-type Salmonella, a pcgL mutant was unable to use D-Ala-D-Ala as a sole carbon source. The pcgL gene conferred D-Ala-D-Ala dipeptidase activity upon Escherichia coli K-12 but did not allow growth on D-Ala-D-Ala. The PcgL protein localizes to the periplasmic space of Salmonella, suggesting that this dipeptidase participates in peptidoglycan metabolism. 相似文献
15.
16.
Angiotensin converting enzyme: implications from molecular biology for its physiological functions. 总被引:1,自引:0,他引:1
N M Hooper 《The International journal of biochemistry》1991,23(7-8):641-647
1. The two isozymes of human angiotensin converting enzyme (ACE; EC 3.4.15.1) have recently been cloned and sequenced. 2. The larger, endothelial isozyme has two highly similar internal domains each bearing a putative catalytic site. In contrast the smaller, testicular isozyme has a single catalytic site corresponding to the C-terminal domain of endothelial ACE and represents the ancestral, non-duplicated form of the gene. 3. Both isozymes are anchored in the plasma membrane by a single hydrophobic transmembrane polypeptide located near the C-terminus, and both are extensively N-glycosylated. 4. The testicular isozyme may also be O-glycosylated. 5. The soluble form of ACE in plasma, seminal fluid and other body fluids appears to be derived from the membrane-bound endothelial isozyme by a post-translational modification. 6. ACE has a complex substrate specificity with peptidyl tripeptidase or endopeptidase action on certain peptides, as well as the classical peptidyl dipeptidase activity. 7. Numerous potent inhibitors of the enzyme have been developed and used successfully in the treatment of hypertension, but some of the observed side effects may be due to inhibition of other zinc metalloenzymes. 8. Both endothelial and testicular ACE are highly conserved between species, indicative of the essential role(s) of the enzyme in blood pressure regulation and other physiological processes. 相似文献
17.
Immunoelectron microscopic demonstration of an esterase on the outer membrane of Xanthomonas maltophilia.
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Xanthomonas maltophilia (later synonym of Pseudomonas maltophilia), an ubiquitous species, is known to show proteolytic and lipolytic activities. A cell-bound esterase which hydrolyzes beta-naphthyl acetate during growth has been extracted from a strain isolated from soil. Because of its strongly hydrophobic character, the enzyme could be efficiently solubilized only by Triton X-100. This nonionic detergent must be added in polyacrylamide gels to permit migration. Polyclonal rabbit antibodies raised against the Triton-soluble esterase complex were used to localize the enzyme at the ultrastructural level. Electron microscopy of cell sections of this organism and immunogold labeling demonstrated that the enzyme was located on the outer membrane. Such an envelope-bound esterase may produce assimilable substrates for X. maltophilia which can grow in various environments. 相似文献
18.
Xanthomonas maltophilia (later synonym of Pseudomonas maltophilia), an ubiquitous species, is known to show proteolytic and lipolytic activities. A cell-bound esterase which hydrolyzes beta-naphthyl acetate during growth has been extracted from a strain isolated from soil. Because of its strongly hydrophobic character, the enzyme could be efficiently solubilized only by Triton X-100. This nonionic detergent must be added in polyacrylamide gels to permit migration. Polyclonal rabbit antibodies raised against the Triton-soluble esterase complex were used to localize the enzyme at the ultrastructural level. Electron microscopy of cell sections of this organism and immunogold labeling demonstrated that the enzyme was located on the outer membrane. Such an envelope-bound esterase may produce assimilable substrates for X. maltophilia which can grow in various environments. 相似文献
19.
Bradykinin (BK) (Arg1-Pro2-Pro3-Gly4-Phe5-Ser6-Pro7-Phe8-Arg9) was degraded by rat brain synaptic membranes at a rate comparable to that found for Met-enkephalin, but approximately 40 times the rate for vasopressin and oxytocin. The catabolic pathway for BK and its metabolites was elucidated through the use of high performance liquid chromatography for metabolite identification and peptidase inhibitors for blocking specific cleavage sites. BK was hydrolyzed at three sites: at the -Phe5-Ser6- bond by metalloendopeptidase 24.15, at the -Pro7-Phe8- bond by an apparently novel peptidyl dipeptidase, and at the -Phe8-Arg9 bond by a carboxypeptidase B-like enzyme. Each enzyme contributed about equally to BK degradation under the assay conditions used. Some of the resulting metabolites were further hydrolyzed: BK(1-8) to BK(1-7) + Phe by a DFP inhibitable prolyl carboxypeptidase-like enzyme, BK(1-8) to BK(1-5) + BK(6-8) by metalloendopeptidase 24.15, BK(1-7) slowly to BK(1-5) by a second peptidyl dipeptidase which was captopril inhibited, and Phe-Arg to Phe + Arg by a bestatin-inhibited dipeptidase. A number of properties of the individual enzymes were determined including sensitivity to a variety of peptidase inhibitors. These results provide a starting point for investigating the potential physiological role of each enzyme in BK function in the brain. 相似文献
20.
The dipeptidase activity of an unpurified soluble extract of the cotyledons of Cucurbita maxima Duch. var. Hubbard remained unchanged during the first 2 days of germination and then increased at a rapid rate during the next 3 days. The dipeptidase activity of two of three lots of seeds required the presence of the embryo axis for maximal dipeptidase activity, whereas the third lot was uninfluenced by the embryo axis. This discrepancy was possibly due to genetic differences. In those seeds which required the presence of the embryo axis for maximal dipeptidase activity, the cytokinin benzyladenine could replace the embryonic axis. When the protein synthesis inhibitor cycloheximide was added to the seeds at the beginning of germination, it inhibited dipeptidase activity of the cotyledons from 26 to 55 per cent, depending of the basis of calculation, at 5 days. When the cycloheximide was added to 3-day-old seedling the inhibition of dipeptidase activity in the cotyledons was almost immediate. The relative hydrolysis of
-leucylglycine and glycylglycine were compared after temperature inactivation and purification; the results showed that more than one enzyme was responsible for the dipeptidase activity. The presence of a dialysable dipeptidase inhibitor(s) was demonstrated. Relatively high dipeptidase activity was also found in the roots and shoots. 相似文献