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91.
Purification and characterization of a methionine-specific aminopeptidase from Salmonella typhimurium 总被引:1,自引:0,他引:1
P Wingfield P Graber G Turcatti N R Movva M Pelletier S Craig K Rose C G Miller 《European journal of biochemistry》1989,180(1):23-32
An aminopeptidase specific for methionine (peptidase M) has been purified from wild-type and mutant Salmonella typhimurium strains. Recombinant peptidase M was also purified from Escherichia coli. These preparations were characterized with respect to their physicochemical properties using analytical ultracentrifugation, SDS/PAGE, isoelectric focusing, titration curve analysis, amino acid analysis, N-and C-terminal sequencing and various spectroscopic methods. Peptidase M activity is stimulated by Co2+, in agreement with previous studies using crude extracts of Salmonella. The purified preparations did not contain significant amounts of any metal. Enzymically important metal is loosely associated and lost during enzyme purification. Peptidase M was shown to contain seven free sulphydryl residues none of which are involved in either intra-or inter-molecular disulphide bonds. Most appear solvent-accessible as evidenced by their reactivity under native conditions. Limited modification of the sulphydryl residues with either iodoacetamide or 5,5'-dithiobis(2-nitrobenzoic acid) led to inactivation. Several cysteines were shown to be labelled to various degrees by peptide mapping of inactivated S-[14C]carboxymethylated protein. Whether cysteine modification affects enzymic activity directly (blocking an active site) or indirectly (by causing conformational change) remains to be established. 相似文献
92.
D W Reif V M Samokyszyn D M Miller S D Aust 《Archives of biochemistry and biophysics》1989,269(2):407-414
The diabetogenic action of alloxan is believed to involve oxygen free radicals and iron. Incubation of glutathione (GSH) and alloxan with rat liver ferritin resulted in release of ferrous iron as assayed by spectrophotometric detection of ferrous-bathophenanthroline complex formation. Neither GSH nor alloxan alone mediated iron release from ferritin. Superoxide dismutase (SOD) and catalase did not affect initial rates of iron release whereas ceruloplasmin was an effective inhibitor of iron release. The reaction of GSH with alloxan resulted in the formation of the alloxan radical which was detected by ESR spectroscopy and by following the increase in absorbance at 310nm. In both instances, the addition of ferritin resulted in diminished alloxan radical detection. Incubation of GSH, alloxan, and ferritin with phospholipid liposomes also resulted in lipid peroxidation. Lipid peroxidation did not occur in the absence of ferritin. The rates of lipid peroxidation were not affected by the addition of SOD or catalase, but were inhibited by ceruloplasmin. These results suggest that the alloxan radical releases iron from ferritin and indicates that ferritin iron may be involved in alloxan-promoted lipid peroxidation. 相似文献
93.
Amino acid sequence of a mouse mucosal mast cell protease 总被引:11,自引:0,他引:11
H L Trong G F Newlands H R Miller H Charbonneau H Neurath R G Woodbury 《Biochemistry》1989,28(1):391-395
The amino acid sequence has been determined of a mouse mucosal mast cell protease isolated from the small intestines of mice infected with Trichinella spiralis. The active protease contains 226 residues. Those corresponding to the catalytic triad of the active site of mammalian serine proteases (His-57, Asp-102, and Ser-195 in chymotrypsin) occur in identical positions. A computer search for homology indicates 74.3% and 74.1% sequence identity of the mouse mast cell protease compared to those of rat mast cell proteases I and II (RMCP I and II), respectively. The six half-cystine residues in the mouse mast cell protease are located in the same positions as in the rat mast cell proteases, cathepsin G, and the lymphocyte proteases, suggesting that they all have identical disulfide bond arrangements. At physiological pH, the mouse and rat mucosal mast cell proteases have net charges of +3 and +4, respectively, as compared to +18 for the protease (RMCP I) from rat connective tissue mast cells. This observation is consistent with the difference in solubility between the mucosal and connective tissue mast cell proteases when the enzymes are extracted from their granules under physiological conditions. 相似文献
94.
Affinity-labeling probes with sites of cross-linking distributed along the ligand have been used to biochemically characterize the pancreatic cholecystokinin (CCK) receptor. Probes with photolabile sites spanning the receptor-binding domain have labeled a Mr = 85,000-95,000 plasma membrane protein, while a probe cross-linked via the amino terminus of CCK-33, far removed from the carboxyl-terminal receptor-binding domain, has labeled a distinct Mr = 80,000 protein. In this work, protease peptide mapping of the pancreatic proteins labeled by each of these probes has been performed to gain insight into the identities of the bands and to define domains of the labeled proteins. Photolabile decapeptide probes with sites of cross-linking at the amino terminus, mid region, and carboxyl terminus of the receptor-binding domain each labeled a Mr = 85,000-95,000 glycoprotein with a Mr = 42,000 core protein and similar Staphylococcus aureus V8 protease peptide maps. This confirms that each probe labels the same binding protein and the same domain of that protein. Serial slices through the broad labeled band were separately deglycosylated and protease-treated, demonstrating a single protein core with differential glycosylation. The CCK-33-based probe, however, labeled predominantly two proteins, one having similar sizes in its native and deglycosylated forms to that labeled by the decapeptide probes and a distinct Mr = 80,000 protein. Of note, the peptide map of the protein believed to be the same as that labeled by the shorter probes was different, suggesting that this probe labeled the binding subunit at a site distinct from that which was labeled by the short probes. 相似文献
95.
Role of surface electrostatics in the operation of a high-conductance Ca2+-activated K+ channel 总被引:8,自引:0,他引:8
This paper demonstrates that local electric fields originating from negatively charged groups on a K+-specific ion channel modify its behavior. Single high-conductance, Ca2+-activated K+ channels were studied in neutral phospholipid bilayers. The channel protein surface charges were manipulated experimentally by carboxyl group esterification using trimethyloxonium (TMO) or by electrolyte screening. Three channel properties--ion conduction, ion blockade, and voltage-dependent gating--are affected by surface electrostatics. Negative charges increase the affinity of cationic pore blockers by establishing a local negative potential at the pore entrance; these charges modify channel gating by establishing a potential gradient across the ion channel; finally, both effects influence ion permeation through the pore. 相似文献
96.
Dehydroisoandrosterone prevention of autoimmune disease in NZB/W F1 mice: lack of an effect on associated immunological abnormalities 总被引:1,自引:0,他引:1
The effect of dietary dehydroisoandrosterone (DHA) on several immunological abnormalities associated with the development of systemic lupus erythematosus in New Zealand Black/New Zealand White F1 (NZB/W) female mice was examined. Despite the extraordinary benefits in prolonged survival and decreased synthesis of antibodies to double-stranded DNA obtained by adding DHA (0.4% w/v) to the diet fed to these mice (Lucas et al. (1985) J. Clin. Invest. 75, 2091-2093), remarkably small changes in the chemistry and function of the immune system were detected. DHA prevented the increases in spleen mass and in peritoneal cell number which occur with age in NZB/W female mice, but did not prevent the development of hypergammaglobulinemia. DHA did not affect peritoneal macrophage functions as measured by the phagocytosis of opsonized and non-opsonized sheep erythrocytes, or the zymosan-stimulated release of PGE2, 6-ketoPGF1 alpha, TXB2 and LTC4. In spleen, DHA delayed the loss of T-cell mitogenic responses until 5.5 months of age, but did not alter the spleen lymphocyte population. 相似文献
97.
This study provides the most comprehensive physico-chemical and phytoplankton data yet available for Australian dune lakes,
which are among the world's most naturally acidic and oligotrophic freshwaters. Seasonal and spatial variations were examined
in Blue Lagoon and Lake Freshwater, two ‘water-table window’ lakes in south-east Queensland. Like other dune lakes, they are
acidic (minimum pH 4.20 and 4.55, respectively), polymictic water bodies with low concentrations of marine-derived major ions
and almost undetectable levels of trace metals. While linmologically similar in winter, during spring-summer Lake Freshwater
has significantly higher levels of chlorophyll-a, total phosphorus (TP) and turbidity than Blue Lagoon and other dune lakes, indicating seasonal mesotrophy. The key nutrient
is TP, which has recently increased to a maximum of 17 μ· l−1, due either to inputs from recreational sources, or to the death and decomposition of littoral vegetation resulting from
falling water levels over the last decade. Inorganic nitrogen, though present only in small amounts, does not appear to limit
the eutrophication process because of a shift in phytoplankton dominance from the usual desmids and dinoflagellates to N2-fixing blue-green algae.
A chlorophyll - TP linear regression derived for dune lakes indicates that at TP < 20 μg · l−1 chlorophyll ‘yield’ is higher than in other lake types represented by regressions from the literature. This may be due to
a more efficient utilization of the limited available phosphorus by dune lake algae which have adapted to the naturally oligotrophic
environment. The implications of these findings for lake management are discussed. 相似文献
98.
Assignment of cathepsin E (CTSE) to human chromosome region 1q31 by in situ hybridization and analysis of somatic cell hybrids 总被引:2,自引:0,他引:2
J M Couvreur T Azuma D A Miller M Rocchi T K Mohandas F A Boudi R T Taggart 《Cytogenetics and cell genetics》1990,53(2-3):137-139
A full length cathepsin E (CTSE) cDNA clone was used to assign the corresponding gene to human chromosome region 1q31 by in situ hybridization. Southern blot analysis of DNA from three independent human x rodent somatic cell hybrids containing X;1 translocations confirmed the assignment of the CTSE gene to the distal region of the long arm of chromosome 1. 相似文献
99.
Marc E. Savard J. David Miller Baharuddin Salleh Richard N. Strange 《Mycopathologia》1990,110(3):177-181
Extracts of rice on which an isolate of Fusarium chlamydosporum had been cultured were toxic to brine shrimps. The toxic fraction was purified by flash chromatography to give two compounds which were identified by UV, IR, NMR and mass spectroscopy at the 6 and 6 isomers of 5-hydroxy-4-methoxy-6, 8a-dimethyl-6,7-dihydro-2H,8aH-pyrano[2,3-b]pyran-2-one. These lactones for which the name chlamydosporol is proposed have not been reported previously. When tested in brine shrimp and HeLa cell assays, the LC50 concentration for a mixture of the isomers was approximately 400 g/ml in both systems. 相似文献
100.
Summary The postulate that 60-Hz electric field-induced bioeffect severity is proportional to induced transmembrane potential [
] magnitude was tested and supported using a plant root model cell system. Statistically significant correlations were obtained upon regression of the relative rates of exposedVicia faba andZea mays root segment growth on the average
(calculated) arising in those segments under specified 60 Hz field exposure conditions. The
associated with the apparent threshold for growth inhibition was similar inZea andVicia roots (2.5 vs 2.4 mV, respectively). At
greater than this threshold,Zea root growth declined by about 9% per mV, andVicia root growth by about 19% per mV induced potential.Abbreviations EF
electric field
- RGR
relative growth rate
- RSGR
relative segmental growth rate
-
induced membrane potential
-
segmental-average induced membrane potential
-
VC
d
region of root tip in which a complete, nascent vascular cylinder is first distinguishable in histological sections 相似文献