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1.
Single-site chromosomal Tn5 insertions affect the export of pectolytic and cellulolytic enzymes in Erwinia chrysanthemi EC16 总被引:9,自引:0,他引:9
Exponentially growing cells of Erwinia chrysanthemi EC16 usually export about 98% of their pectate lyase (PL) and protease, about 40% of their polygalacturonase (PG), and about 60% of their cellulase (endoglucanase or carboxymethyl cellulase; CL). By using the R plasmid, pJB4JI (pPH1JI::Mu::Tn5), three independent Tn5 insertion mutants were obtained that exported normal levels of protease but 10% or less of PL, PG, and CL. Physical analysis revealed that single copies of Tn5 had inserted into the E. chrysanthemi chromosome, producing a similar export-defective (Out-) phenotype. The synthesis of PL, PG, and CL was not affected by the Tn5 insertions. These enzymes were released from the mutants on spheroplast formation, indicating that they were located in the periplasmic space. Tn5 insertions caused the loss of a 35-kilodalton periplasmic protein, but did not alter the outer membrane protein composition. The findings are discussed with respect to the current knowledge on protein export in gram-negative bacteria. 相似文献
2.
Homogenates of trophozoites of Entamoeba histolytica released glucose 1-phosphate from amylopectin, glycogen, and amylose in a ratio of 100:78:74 at glucopolysaccharide concentrations of 0.1%. By use of self-generating Percoll gradients this activity was shown to be particulate and associated with glycogen. The phosphorylase was extracted from the 40,000 g pellet in aqueous medium and purified to homogeneity by gel filtration on Fractogel TSK HW-55(F) followed by chromatography on Blue Sepharose CL-6B. The purified enzyme was active not only against the glucopolysaccharides but also on dextrins with more than 3 glucose moieties, which were primarily formed by the action of amoebic amylases. At substrate concentrations of 1 mM nonreducing ends of each glucan, the phosphorolysis rate of the branched polysaccharides was about 1.75 x 10(4) times higher than those of the maltodextrins. By means of HPLC the sequential degradation of 4-nitrophenyl-maltoheptaoside (G(7)-pNP) was studied. Native phosphorylase exhibited a relative molecular mass of M(r) = 200,000 by gel filtration and gel electrophoresis. The SDS electrophoresis, under reducing conditions, indicated that the native enzyme was a dimer. Optimal degradation of the polysaccharides and dextrins was achieved at pH values of 7.5 and 7.0 respectively. 相似文献
3.
Ponnada Suresh Kumar KK Pulicherla Mrinmoy Ghosh Anmol Kumar KRS Sambasiva Rao 《Bioinformation》2011,6(8):311-314
Enzymes from psychrophiles catalyze the reactions at low temperatures with higher specific activity. Among all the psychrophilic enzymes produced, cold active
β-galactosidase from marine psychrophiles revalorizes a new arena in numerous areas at industrial level. The hydrolysis of lactose in to glucose and galactose by
cold active β-galactosidase offers a new promising approach in removal of lactose from milk to overcome the problem of lactose intolerance. Herein we propose, a
3D structure of cold active β-galactosidase enzyme sourced from Pseudoalteromonas haloplanktis by using Modeler 9v8 and best model was developed having
88% of favourable region in ramachandran plot. Modelling was followed by docking studies with the help of Auto dock 4.0 against the three substrates lactose,
ONPG and PNPG. In addition, comparative docking studies were also performed for the 3D model of psychrophilic β-galactosidase with mesophilic and
thermophilic enzymes. Docking studies revealed that binding affinity of enzyme towards the three different substrates is more for psychrophilic enzyme when
compared with mesophilic and thermophilic enzymes. It indicates that the enzyme has high specific activity at low temperature when compared with mesophilic
and thermophilic enzymes. 相似文献
4.
Melanoma is the most lethal cutaneous cancer with a highly aggressive and metastatic phenotype. While recent genetic and epigenetic studies have shed new insights into the mechanism of melanoma development, the involvement of regulatory non‐coding RNAs remain unclear. Long non‐coding RNAs (lncRNAs) are a group of endogenous non‐protein‐coding RNAs with the capacity to regulate gene expression at multiple levels. Recent evidences have shown that lncRNAs can regulate many cellular processes, such as cell proliferation, differentiation, migration and invasion. In the melanoma, deregulation of a number of lncRNAs, such as HOTAIR, MALAT1, BANCR, ANRIL, SPRY‐IT1 and SAMMSON, have been reported. Our review summarizes the functional role of lncRNAs in melanoma and their potential clinical application for diagnosis, prognostication and treatment. 相似文献
5.
Changes in odor quality discrimination following recovery from olfactory nerve transection 总被引:4,自引:2,他引:2
Following recovery from olfactory nerve transection, animals regain their
ability to discriminate between odors. Odor discrimination is restored
after new neurons establish connections with the olfactory bulb. However,
it is not known if the new connections alter odor quality perception. To
address this question, 20 adult hamsters were first trained to discriminate
between cinnamon and strawberry odors. After reaching criterion (> or =
90% correct response), half of the animals received a bilateral nerve
transection (BTX) and half a surgical sham procedure. Animals were not
tested again until day 40, a point in recovery when connections are
re-established with the bulb. When BTX animals were tested without food
reinforcement, they could not perform the odor discrimination task. Sham
animals, however, could discriminate, demonstrating that the behavioral
response had not been extinguished during the 40 day period. When
reinforcement was resumed, BTX animals were able to discriminate between
cinnamon and strawberry after four test sessions. In addition, their
ability to discriminate between these two familiar odors was no different
than that of BTX and sham animals tested with two novel odors, baby powder
and coffee. These findings suggest that, after recovery from nerve
transection, there are alterations in sensory perception and that
restoration of odor quality discrimination requires that the animal must
again learn to associate individual odor sensations with a behavioral
response.
相似文献
6.
Single-site chromosomal Tn5 insertions affect the export of pectolytic and cellulolytic enzymes in Erwinia chrysanthemi EC16. 总被引:11,自引:1,他引:10 下载免费PDF全文
Exponentially growing cells of Erwinia chrysanthemi EC16 usually export about 98% of their pectate lyase (PL) and protease, about 40% of their polygalacturonase (PG), and about 60% of their cellulase (endoglucanase or carboxymethyl cellulase; CL). By using the R plasmid, pJB4JI (pPH1JI::Mu::Tn5), three independent Tn5 insertion mutants were obtained that exported normal levels of protease but 10% or less of PL, PG, and CL. Physical analysis revealed that single copies of Tn5 had inserted into the E. chrysanthemi chromosome, producing a similar export-defective (Out-) phenotype. The synthesis of PL, PG, and CL was not affected by the Tn5 insertions. These enzymes were released from the mutants on spheroplast formation, indicating that they were located in the periplasmic space. Tn5 insertions caused the loss of a 35-kilodalton periplasmic protein, but did not alter the outer membrane protein composition. The findings are discussed with respect to the current knowledge on protein export in gram-negative bacteria. 相似文献
7.
pULB113, an RP4::mini-Mu plasmid, mediates chromosomal mobilization and R-prime formation in Erwinia amylovora, Erwinia chrysanthemi, and subspecies of Erwinia carotovora. 总被引:1,自引:0,他引:1 下载免费PDF全文
The RP4::mini-Mu plasmid pULB113, transferred from Escherichia coli strain MXR, was stable and transfer proficient in Erwinia amylovora strain EA303, E. carotovora subsp. atroseptica strain ECA12, E. carotovora subsp. carotovora strain ECC193, and E. chrysanthemi strain EC183. The plasmid mobilized an array of Erwinia sp. chromosomal markers (E. amylovora: his+,ilv+,rbs+,ser+,thr+;E. chrysanthemi:arg+,his+,ilv+,leu+; E. carotovora subsp. atroseptica: arg+,gua+,leu+,lys+,pur+,trp+; E. carotovora subsp. carotovora: arg+,gua+,leu+,lys+,out+[export of enzymes],pur+,trp+), suggesting random interactions of the plasmid with the chromosomes. In E. carotovora subsp. carotovora, pULB113-mediated two-factor crosses revealed linkage between three auxotrophic markers and the out loci. The export of pectate lyase, polygalacturonase, and cellulase and the maceration of potato tuber tissue occurred with Out+, but not Out-, strains of E. carotovora subsp. carotovora, indicating the importance of enzyme export in plant tissue maceration. Erwinia sp. donors harboring pULB113 complemented mutations in various biosynthetic and catabolic genes (arg, gal, his, leu, met, pro, pur, thy) in Escherichia coli recA strains. Escherichia coli transconjugants harbored pULB113 primes as indicated by the cotransfer of Erwinia genes and pULB113 markers and a change in plasmid mass. Moreover, the PstI and SmaI cleavage patterns of selected pULB113 primes were different from those of pULB113. pULB113 primes carried DNA insertions ranging from 3 to about 160 kilobases. These findings indicate that pULB113 is useful for in vivo gene cloning and genetic analysis of various enterobacterial phytopathogens. 相似文献
8.
Isolation and characterization of Tn5 insertion mutants of Erwinia chrysanthemi that are deficient in polygalacturonate catabolic enzymes oligogalacturonate lyase and 3-deoxy-D-glycero-2,5-hexodiulosonate dehydrogenase. 总被引:10,自引:6,他引:4 下载免费PDF全文
Mutants of Erwinia chrysanthemi EC16 deficient in the polygalacturonate catabolic enzymes oligogalacturonate lyase (Ogl-) and 3-deoxy-D-glycero-2,5-hexodiulosonate (ketodeoxyuronate) dehydrogenase (KduD-) were obtained by Tn5 mutagenesis using the R plasmid pJB4JI. Ogl- Exu+ (Exu+, D-galacturonate utilization) and KduD- Exu- strains macerated potato tuber tissue and utilized glucose, glycerol, and gluconate, but they did not utilize polygalacturonate, unsaturated digalacturonate, or saturated digalacturonate. Genetic and physical evidence indicated that the Ogl- mutants and a KduD- recombinant contained a single copy of Tn5 and that Tn5 (Kmr) was linked to the mutant phenotypes. In the Ogl+ parents, basal levels of oligogalacturonate lyase were present in glycerol-grown cells and induced levels were present with saturated or unsaturated digalacturonate, while oligogalacturonate lyase was undetectable under similar conditions in Ogl- strains. Pectate lyase, polygalacturonase, and ketodeoxyuronate dehydrogenase were induced in an Ogl- strain by 3-deoxy-D-glycero-2,5-hexodiulosonate and by the enzymatic products of unsaturated digalacturonate but not by the digalacturonates. The KduD- strains lacked the dehydrogenase activity but in the presence of the digalacturonates produced higher levels of pectate lyase, polygalacturonase, and oligogalacturonate lyase than the KduD+ parents did. In the KduD- strains, pectate lyase and oligogalacturonate lyase were induced by unsaturated digalacturonate in a "gratuitous" manner, suggesting an intracellular accumulation of the inducer(s). We conclude that an intermediate(s) of the ketodeoxyuronate pathway induces pectate lyase, polygalacturonase, oligogalacturonate lyase, and ketodeoxyuronate dehydrogenase in E. chrysanthemi. 相似文献
9.
Preincubation of a triolein/phospholipid/cholesteryl oleate-emulsion in vitro with either pancreatic phospholipase A2 (PLA2) or gastric lipase (GL) resulted in hydrolysis (measured by pH-stat-titration) of cholesteryl [3H]oleate only after human pancreatic carboxyl ester lipase (CEL) was added to the system. No appreciable hydrolysis was observed when CEL was added alone. Consequently, a concerted action either of PLA2 and CEL or of GL and CEL made the substrate cholesteryl oleate available for hydrolysis by CEL. This was the case when cholesteryl oleate was solubilised in a phospholipid-stabilised triglyceride emulsion, which is the physico-chemical form in which the major part of dietary cholesteryl esters are presented to the gastro-intestinal tract of man. 相似文献
10.
Douglas G. Ward Laura Baxter Naheema S. Gordon Sascha Ott Richard S. Savage Andrew D. Beggs Jonathan D. James Jennifer Lickiss Shaun Green Yvonne Wallis Wenbin Wei Nicholas D. James Maurice P. Zeegers KK Cheng Glenn M. Mathews Prashant Patel Michael Griffiths Richard T. Bryan 《PloS one》2016,11(2)