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1.
The effect of a decrease inpH on the structural integrity of carmin has been monitored by a variety of biophysical techniques. The protein undergoes initial dissociation up topH 3.5–4.0 without any significant denaturation. Below thispH the process of dissociation and denaturation appears to be simultaneous. Further, in thepH range of 2.5–1.6 the protein reassociates to probably a different polymer resulting from possibly, an entropically driven hydrophobic interaction. The process of dissociation appears to be reversible to a large extent. The process of denaturation appears to be governed by the kinetic path that the denatured protein molecule follows either by a sudden decrease inpH or through a gradual decrease inpH. These results are interpreted while keeping in view the oligomeric and globular structure of carmin at neutralpH. The results would help in understanding of structure-function relationship of the protein and its role in hydrogen ion bindingin vivo. 相似文献
2.
Uri S. Ladror Gary T. Wang William L. Klein Thomas F. Holzman Grant A. Krafft 《The protein journal》1994,13(4):357-366
Fluorogenic peptide substrates designed to encompass the reportedα-secretory and amyloidogenic cleavage sites of the amyloid-β precursor protein (βPP) were used to analyze proteinase activities in brain extracts from control patients and those with Alzheimer's disease (AD). Activity against the secretory substrate atpH 7.5 in control and AD brains produced a major endopeptidase cleavage at the Lys687-Leu688 bond (βPP770 numbering), consistent with theβPP secretase cleavage. Activity in control brains against the amyloidogenic substrate atpH 7.5 produced one cleavage at the Ala673-Glu674 bond, two residues C-terminal to the amyloidogenic Met-Asp site. However, in three of four AD brains, the major cleavage was at the Asp-Ala bond, one residue from the amyloidogenic site. Both endopeptidase and carboxypeptidase activities in AD brains were lower than in control brains. Proteinase activities against the secretory substrate had a major optimum atpH 3.0–4.0 and another atpH 6.0–7.5. Proteinase activities against the amyloidogenic substrate had a major optimum at or belowpH 3.0 and another atpH 6.0. Using both substrates, activities at lowpH were higher in AD brains than in controls, while atpH above 6.5, activities in control brains were higher than in AD. These results indicate that the levels of proteolytic enzymes in AD brains are altered relative to controls. 相似文献
3.
James L. Kerwin 《Journal of invertebrate pathology》1983,41(2):233-237
Zygotes of the obligately parasitic watermold Coelomomyces psorophorae encyst on the cuticular surface of susceptible mosquito larvae as the initial step in the infection process. Zygotes can successfully encyst on Culiseta inornata larvae at temperatures from 12°–30°C, with an optimum of 25°C. Optimum encystment on larvae occurs from pH 6.5–8.5, with activity documented from pH 5.0–9.5. Zygotes are active in water with total salinity of 5.4 ppt down to double glass-distilled water. 相似文献
4.
《Biochimica et Biophysica Acta - Proteins and Proteomics》2018,1866(12):1242-1248
Interaction with model phospholipid membranes of lupin seed γ-conglutin, a glycaemia-lowering protein from Lupinus albus seeds, has been studied by means of Fourier-Transform infrared spectroscopy at p2H 7.0 and at p2H 4.5. The protein maintains the same secondary structure both at p2H 7.0 and at p2H 4.5, but at p2H 7.0 a higher 1H/2H exchange was observed, indicating a greater solvent accessibility. The difference in Tm and TD1/2 of the protein at the abovementioned p2H's has been calculated around 20 °C. Infrared measurements have been then performed in the presence of DMPG and DOPA at p2H 4.5. DMPG showed a little destabilizing effect while DOPA exerted a great stabilizing effect, increasing the Tm of γ-conglutin at p2H 4.5 of more than 20 °C. Since γ-conglutin at p2H 4.5 is in the monomeric form, the interaction with DOPA likely promotes the oligomerization even at p2H 4.5. Interaction between DMPG or DOPA and γ-conglutin has been confirmed by turbidity experiments with DMPC:DMPG or DOPC:DOPA SUVs. Turbidity data also showed high-affinity binding of γ-conglutin to anionic SUVs made up with DOPA. The molecular features outlined in this study are relevant to address the applicative exploitation and to delineate a deeper comprehension of the natural functional role of γ-conglutin. 相似文献
5.
The uptake of iron and the formation of penicillinase was examined in cultures of wild-type Staphylococcus aureus. Uptake of iron was about twice as great at pH 4.7 as at pH 7.4 At pH 4.7, increase in iron uptake in the range of 1.0 to 4.0 μg per mg of bacterial protein was associated with a progressive increase in the rate of penicillinase formation, but a direct correlation between cellular iron content and rate of enzyme formation was not demonstrated. Addition of iron to deferrated medium enhanced penicillinase formation at pH 6.5 to 7.4 two- to fourfold in cultures induced with benzylpenicillin and in uninduced cultures. To demonstrate an effect on the uninduced cells, it was necessary to increase iron uptake by preliminary incubation of cells with iron in buffer. Calcium and certain other ions depressed iron uptake at acidic and at neutral pH, and, presumably as a result of this action, depressed the formation of penicillinase. Iron did not enhance penicillinase formation at pH 4.7 by two penicillinase constitutive mutants nor by wild-type cells undergoing induction at pH 6.5 by cephalosporin C or methicillin. After removal of cephalosporin C or methicillin during an early phase of induction, residual synthesis of enzyme was increased by prior uptake of iron. The results are considered compatible with the concept that uptake of iron, especially at acidic pH, interferes with the formation or function of penicillinase repressor. 相似文献
6.
Type E botulinum neurotoxin is produced byClostridium botulinum along with a neurotoxin binding protein which helps protect the neurotoxin from adversepH, temperature, and proteolytic conditions. The neurotoxin binding protein has been purified as a 118-kDa protein. Secondary structure content of the neurotoxin binding protein as revealed by far-UV circular dichroism spectroscopy was 19% α-helix, 50%β-sheets, 28% random coils, and 3%β-turns. This compared to 22% α-helix, 44%β-sheets, 34% random coils, and noβ-turns of the type E botulinum neurotoxin. The complex of the two proteins revealed 25%α-helix, 45%β-sheets, 27% random coils, and 3%β-turns, suggesting a significant alteration at least in theα-helical folding of the two proteins upon their interaction. Tyrosine topography is altered considerably (28%) when the neurotoxin and its binding protein are separated, indicating strong interaction between the two proteins. Gel filtration results suggested that type E neurotoxin binding protein clearly complexes with type E neurotoxin. The interaction is favored at lowpH as indicated by an initial binding rate of 8.4 min?1 atpH 5.7 compared to 4.0 min?1 atpH 7.5 as determined using a fiber optic-based biosensor. The neurotoxin and its binding protein apparently are of equivalent antigenicity, as both reacted equally on enzyme-linked immunosorbent assay to polyclonal antibodies raised against the toxoid of their complex. 相似文献
7.
Junko Nishiitsutsuji-Uwo Ayako Ohsawa Masaji S. Nishimura 《Journal of invertebrate pathology》1977,29(2):162-169
The stability and solubility of the crystal toxin of Bacillus thuringiensis var. aizawai were measured using larvae of Bombyx mori as the test animals under the same test conditions. Insecticidal activity was retained in buffer solutions of pH 2.2–10.8. It decreased rapidly either below pH 2.0 or above pH 11. As to the effects of time and temperature, the longer the incubation time, the lower was the activity in solutions above pH 11, and the higher the treatment temperature, the lower was the pH at which degradation of activity occurred. Guanidine (6 m) with 1% mercaptoethanol was not good for dissolving the toxic protein. Instead, 8 m urea with 0.5% dithiothreitol or with 1% mercaptoethanol and 0.04 m NaOH seemed better, and onefourth of the total activity appeared in the soluble fraction. On the other hand, 0.2 m thioglycollate and “Ellis's” buffer dissolved the toxic moiety completely and safely. 相似文献
8.
Effects of pH and ligand binding upon the conformation of Cellobiohydrolase I (CBHI) and endoglucanase I (EGI) from Trichoderma pseudokiningii S-38 have been studied by circular dichroism measurements. In the high-pH range (6–9), increasing pH resulted in a similar conformational change occurring in free CBHI and EGI, while such treatment gave different changes of the two enzyme conformations in the presence of cellobiose. On the other hand, in the low-pH region, with both CBHI an EGI in the active form, decreasing pH resulted in a large conformational change of free EGI compared to that of free CBHI, whereas ligand binding resulted in a similar change of both CBHI and EGI, independent of pH change. 相似文献
9.
Ferredoxin isolated from Halobacterium of the Dead Sea (HFd) was found to be stable and retain its conformation in 4–0.5 M salt solutions. Reconstitution of the denatured protein to the oxidized form in 2H2O indicated that the resonances shifted to the 8–10 ppm region, which include 18 protons, are nonexchangeable -NH protons. The C2H and C4H resonances of His-119 were assigned in both oxidized and reduced HFd. pH titration curves of these resonances yielded a pKa for this His of 6.57 ± 0.1 and 6.65 ± 0.1 in oxidized and reduced HFd, respectively. pH titration curves, T1 relaxation times, and the temperature dependence of the chemical shift were obtained for resonances between 6 and 10 ppm of oxidized HFd. In oxidized HFd a paramagnetically shifted resonance was observed at 15 ppm with 1 H intensity, and an anti-Curie temperature dependence. In reduced HFd eight resonances each with 1 H intensity were shifted downfield by 10–50 ppm and one resonance with 1 H intensity was shifted upfield to ?6.8 ppm. Four of these resonances exhibited an anti-Curie temperature dependence, two exhibited a moderate Curie dependence, and three were temperature independent. 相似文献
10.
Effect of pH on Adenine and Amino Acid Uptake During Sporulation in Saccharomyces cerevisiae 总被引:21,自引:14,他引:7
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Dallice Mills 《Journal of bacteriology》1972,112(1):519-526
During the early stages of sporulation in Saccharomyces cerevisiae, the pH of the acetate sporulation medium rises to values of 8.0 or higher. Associated with this rise in pH is a reduced cell permeability to certain precursors of ribonucleic acid (RNA), deoxyribonucleic acid or protein. Uptake of adenine, alanine, and leucine was optimal at pH 5.6 to 6.0, but sporulation was inhibited when the sporulation medium was buffered below pH 7.0. Cellular impermeability can be largely overcome by adjusting the acetate sporulation medium to pH 6.0 for optimal uptake of 14 C-adenine during short pulses without any apparent effect on sporulation. Sporulating cells pulse-labeled 20 min at pH 6.0 incorporated 40 times more 14C-adenine into RNA than sporulating cells pulse-labeled at pH 8.0. This increased incorporation can be attributed to a 100-fold increase in labeled adenosine triphosphate in cells pulse-labeled at pH 6.0 where maximum uptake occurs. 相似文献
11.
Jose Neptuno Rodriguez-Lopez Simon J. George R. N. F. Thorneley 《Journal of biological inorganic chemistry》1998,3(1):44-52
Horseradish peroxidase isoenzyme C (HRPC) mutants were constructed in order to understand the involvement of two key distal heme cavity residues, histidine 42 and arginine 38, in the formation and structure of the carbon monoxide complex of HRPC (carbonyl HRPC). The rates of CO binding to the wild-type glycosylated and non-glycosylated recombinant (HRPC*) ferrous enzymes were essentially identical and exhibited the same pH dependence with pK as at 7.4 and 4.0. Data obtained with the His-42?→?Leu [(H42L)HRPC*)] and Arg-38?→?Leu [(R38L)HRPC*] mutants allowed the pK a at 7.4 in ferrous HRPC to be assigned to His-42. The infra-red and electronic absorption spectra of HRPC-CO, HRPC*-CO, (R38L)HRPC*-CO and (H42L)HRPC*-CO have been investigated over the pH range 3.0–10.0. HRPC*-CO exhibited two ν?(CO) bands at 1934?cm–1 and 1905?cm–1 whose relative intensity changed with pH, showing an acidic and a basic pK a as previously reported for HRPC [IE Holzbaur; AM English, AA Ismail (1996) J Am Chem Soc 118?:?3354–3359]. (H42L)HRPC*-CO and (R38L)HRPC*-CO exhibited single infra-red bands at 1924.2?cm–1 (pH?7.0) and 1941.5?cm–1 (pH?5.0) respectively. Acidic and alkaline pK as were determined from shifts in the infra-red frequencies and by UV-visible spectrophotometry at the Söret maxima. (H42L)HRPC*-CO exhibited a pK a at ~pH?4.0 but no alkaline pK a. (R38L)HRPC*-CO exhibited a single pK a at pH?6.5. Shifts of 2–3?cm–1 in ν?(CO) with (H42L)HRPC*-CO in D2O show that a distal residue is H-bonding to the CO in this variant at both pD?7.5 and 3.9. However, with (R38L)HRPC*-CO, only a small shift of the ν?(CO) band was observed at pD?5.5. The results are consistent with the involvement of Arg-38 in H-bonding to the CO ligand in HRPC and with His-42 modulating the distribution of carbonyl HRPC conformers below pH?8.7. These data are discussed in terms of the importance of distal pocket polarity in HRPC. It is concluded that His-42 can have a pK a between 4.0 and 8.7 depending on its environment and the nature of the distal ligand at position 38. This enables His-42 to carry out multiple functions during the catalytic cycle of HRPC. 相似文献
12.
Isolation and some properties of components of nuclear polyhedra from the cabbage looper, Trichoplusia ni 总被引:1,自引:0,他引:1
Nuclear polyhedra obtained from diseased cabbage looper, Trichoplusia ni, were digested with sodium carbonate-saline buffer, pH 11.0. The dissolved polyhedra formed 3 general zones when subjected to density gradient centrifugation. The slowest sedimenting component (Zone 1) had an ultraviolet absorption curve typical of protein and a sedimentation coefficient of 11 S. Capsids, 310 × 40 nm, were located in Zone 2. Virus particles were found in 1–3 bands (Zone 3); those with envelopes measured 300 × 72 nm, and those without envelopes measured 300 × 33 nm. Virus preparations stained with phosphotungstic acid at pH 7.0 exhibited extensive disruption whereas preparations stained at pH 3.0 did not. Virus particles in the sodium carbonate-saline-digested polyhedra had a sedimentation coefficient of 1228 S. Virus particles isolated by high speed centrifugation had a sedimentation coefficient of 1530 S. 相似文献
13.
Hemolymph composition of fourth instar larvae of an autogenous strain of Culex pipiens was examined to determine the effects of parasitism by a mermithid nematode, Romanomermis culicivorax. Mosquitoes were reared under two different pH regimens: 4.5 and 7.3. Wet and dry weight of infected mosquitoes reared at either pH were significantly lower than controls. The effects of parasitism in the development of C. pipiens were evaluated from paraffin sections of mosquito larvae 2, 4, and 6 days postinfection. At 2 days postinfection, the infected larvae showed no apparent effects of parasitism; at day 4, the fat body tissue was reduced and imaginal disc development was retarded; and at day 6, parasitized mosquitoes were smaller in cross section, fat body tissue was found only in isolated clumps, and there was a complete absence of imaginal discs. Concentrations of total carbohydrates in hemolymph from infected fourth instar mosquitoes reared at pH 7.3 were reduced. Trehalose and glucose were each reduced by more than half. Total α-amino nitrogen was significantly lower in infected mosquitoes reared at pH 7.3. However, total amino acid concentrations for hemolymph from control and infected larvae reared at pH 7.3 were the same. Methionine sulfoxide decreased 63% and proline increased 2.5 times in infected mosquitoes. Hemolymph protein concentrations were reduced 80% in infected mosquitoes reared at both pHs. The number of hemolymph proteins also declined from 35 to 22 during infection. Two host proteins, 82,000 and 158,000 daltons, remained prominent throughout the mermithid infection. 相似文献
14.
Induction and Reversal of Contact Inhibition of Growth by <Emphasis Type="Italic">p</Emphasis>H Modification 总被引:7,自引:0,他引:7
MANY normal mammalian cells in culture are sensitive to a growth-inhibitory effect of cellular interaction, evidenced by an almost complete arrest of macromolecular synthesis and cellular division at relatively low population densities (contact inhibition of growth)1. It has, however, been shown that the environmental pH is an important element in this phenomenon2. In the bicarbonate-buffered media ordinarily used for animal cell cultures, an initial alkalinization to pH 7.8–8.0 as a result of CO2 loss is followed by the metabolic acidification of the medium to pH 6.8–7.0, at a rate determined by the population density and the metabolic activity of the specific cell. If, by the use of appropriate non-volatile buffers3,4, cells are kept at or near the pH optimal for the specific strain (H. E., unpublished work), the early contact inhibition of growth does not develop and the cells achieve population densities as much as two to four times greater than those ordinarily observed2. 相似文献
15.
Chemical Constituents and Hydrogenase Binding in Cell Envelopes of Vibrio succinogenes 总被引:1,自引:1,他引:0
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The particulate hydrogenase of Vibrio succinogenes is solubilized during treatment of cell envelopes at pH 11.0. Alkali-solubilized enzyme requires sulfhydryl compounds for activity. At neutral pH, soluble enzyme is reincorporated into alkalitreated cell envelopes and no longer requires an additional activator. In the present study, cell envelopes prepared by lysing cells with ethylenediaminetetraacetic acid plus lysozyme (EDTA-lysozyme) were used to determine the chemical composition of cell envelopes and derived pH 11.0 soluble and insoluble fractions and to investigate some properties of the binding and activation of alkali-solubilized hydrogenase. Lysis with EDTA-lysozyme resulted in the formation of spheroplast ghosts. The derived cell envelopes contained 61% protein, 3% ash, 23% lipid, and 1% phosphorus. The alkali-treated cell envelopes contained 50% protein, 2% ash, 24% lipid, and 1% phosphorus. The ash from cell envelopes and alkali-treated cell envelopes was rich in iron and phosphorus and also contained calcium, copper, magnesium, sodium, and zinc. Virtually all of the weight of the ashed samples was accounted for by the oxides of these metals. Since the reconstitution of particulate hydrogenase was achieved with pH 11.0 supernatant solution and precipitate, intact mucopeptide is not essential for hydrogenase binding. Release of hydrogenase during EDTA-lysozyme lysis was found to depend upon an apparent structural change which occurs in the membranes during extended storage at −20 C. 相似文献
16.
Methods for the solubilization of an insect granulosis virus capsule from Trichoplusia ni were studied using turbidity measurements. The solubilization in the solvents employed was most strongly influenced by pH. Various alcohols and hydrogen bond cleaving agents also enhanced solubilization. The most effective solvent tested at pH′s close to neutrality was 60% n-propanol saturated with guanidine. HCl. The effectiveness of this solvent was minimal at pH 5.2. It was shown that the time courses of the solubilization were generally composed of three phases. A plot of the third phase levels versus pH indicated a normal distribution of those capsules or capsular fragments in resisting solubilization. Validity of the calculation of rate constants for the dissociation reaction in the second phase was discussed. Logarithm of the rate constant showed a linear relationship with the reciprocal of absolute temperature. The activation energy for the dissociation calculated from this relationship using the Arrhenius equation indicated that hydrophobic and hydrogen binding forces play the major roles for the stability of the crystalline structure of the capsular protein. Inic binding forces were estimated to be of minor importance. 相似文献
17.
Mode of action of pectic enzymes. II. Further purification of exopolygalacturonate lyase and pectinesterase from Clostridium multifermentans 总被引:2,自引:1,他引:1
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Exopolygalacturonate lyase and pectinesterase from Clostridium multifermentans were purified 156-fold and 178-fold, respectively, by gel filtration chromatography on Sephadex G-200. The activities of both enzymes coincided in a single protein peak. Profiles of the two activities also coincided in diethylaminoethyl-cellulose chromatography and zonal centrifugation. These studies indicated that the esterase and the lyase were either complexed or similar molecular species. The former seems more probable because of the relatively high molecular weight. Both activities were most stable at pH 6.0. The esterase was inactivated rapidly at pH 5 or 7. Lyase preparations were freed of pectinesterase activity by heating for 30 min at 38 C and pH 7.0. 相似文献
18.
A novel insecticidal GroEL protein from Xenorhabdus nematophila confers insect resistance in tobacco
Punam Kumari Shashi Kant Shazmira Zaman Gagan Kumar Mahapatro Nirupama Banerjee Neera Bhalla Sarin 《Transgenic research》2014,23(1):99-107
Xenorhabdus nematophila is an entomopathogenic bacteria. It secretes a GroEL homolog, XnGroEL protein, toxic to its larval prey. GroEL belongs to the family of molecular chaperones and is required for proper folding of cellular proteins. Oral ingestion of insecticidal XnGroEL protein is toxic to Helicoverpa armigera, leading to cessation of growth and development of the larvae. In the present study, the insecticidal efficacy of XnGroEL against H. armigera has been evaluated in transgenic tobacco plant expressing the protein. A 1.7-kb gene encoding the 58-kDa XnGroEL protein was incorporated into the tobacco genome via Agrobacterium-mediated transformation. The stable integration of the transgene was confirmed by Southern blot analysis and its expression by RT-PCR and western blot analyses in transgenic plants. The transgenic lines showed healthy growth and were phenotypically normal. Insect bioassays revealed significant reduction of 100 % in the survival of larvae (p < 0.001) and 55–77 % reduction in plant damage (p < 0.05 and p < 0.001) compared to the untransformed and vector control plants. The results demonstrate that XnGroEL is a novel potential candidate for imparting insect resistance against H. armigera in plants. 相似文献
19.
A synergistic factor than enhances the infection of a nuclear polyhedrosis virus in the armyworm, Pseudaletia unipuncta, was isolated from the occlusion body (capsule) of a granulosis virus of the armyworm. Disc electrophoresis indicated that the purified factor was a single homogeneous compound. Chemical identification and amino acid analysis showed that it was a simple protein, with a molecular weight of 152,000–163,000. Proteolytic enzymes did not markedly reduce the activity of the factor. It could be stored at ?20°C or lyophilized. The synergistic factor displayed properties of an enzyme. It enhanced the hydrolysis of p-nitrophenyl esters of fatty acids with an optimum of pH 9.0. The relative hydrolytic activity increased with increase in number of carbon atoms in the fatty-acid chain from 2 to 8 and gradually decreased with the number of carbon atoms from 10 to 18. Copper sulfate markedly and mercuric chloride completely prevented enhancement of the hydrolysis of butyrate. When the synergistic factor was fed to larvae with mercuric chloride, it did not enhance the nuclear polyhedrosis virus. 相似文献
20.
N. C. Burns 《Journal of nematology》1971,3(3):238-245
''Amsoy'' soybeans were grown for 2 months in nonsterilized Jackson silt loam amended to pH 4.0, 6.0, and 8.0. Nematodes were extracted biweekly from soil and roots. The greatest numbers of Pratylenchus alleni colonized soybean roots at pH 6.0. Hoplolaimus galeatus and members of the Tylenchinae-Psilenchinae survived best at pH 6.0, while numbers o f the Dorylaimoidea were greatest at both pH 6.0 and 8.0. The non-stylet nematodes were recovered in greater numbers from pH 8.0 soil. Potassium, manganese, and phenols were highest in soybean plants grown in pH 4.0 soil, the pH at which there were the fewest nematodes. A thicker suberized outer layer o f root tissue occurred in plants grown at pH 4.0. 相似文献