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排序方式: 共有329条查询结果,搜索用时 18 毫秒
1.
Om P. Malhotra Frank Valencic Eric T. Fossel Karl A. Koehler 《Journal of Protein Chemistry》1991,10(1):31-41
Ca2+ titrations of the intrinsic fluorescence of a series of -carboxyglutamic acid (GLA)-deficient bovine prothrombin fragments 1 yield response Hill plot parameters useful for characterization of the metal ion-binding process. 11-, 10-, and 9-GLA fragments 1 exhibitT
m
(the (Ca2+)total concentration at which ln (B/F)=0 in the response Hill plot) values between 0.2 and 0.3 mM. A 22-fold increase inT
m
to 5.4 mM is observed for 8-GLA fragment 1.T
m
decreases to 3.8 mM for the 7- and 6-GLA proteins. The value ofh, about 2.8±0.2 for 11-, 10-, and 9-GLA fragments 1, abruptly decreases to 1.2–1.3 for 8-, 7-, and 6-GLA fragments 1. The observed degree of quenching induced by saturating levels of calcium ions is affected by both changes in the intrinsic fluorescence of the metal ion-free proteins and in the maximum possible degree of quenching in the presence of calcium. The kinetic characteristics of the calcium ion-induced quenching of the intrinsic fluorescence of 6-GLA fragment 1 are identical to those observed in 10-GLA fragment 1, suggesting that the fluorescence quenching observed in the 6- and 10-GLA fragments 1, while different in magnitude, involves similar processes. Observation of an abrupt change in the relative electrophoretic mobilities of 11- to 9-GLA fragments 1 compared to 8- to 6-GLA fragments 1, in the absence or presence of Ca2+, suggests the existence of a major protein conformation change which occurs concomitantly with the noted changes inT
m
andh response Hill plot parameters. Molecular mechanics calculations suggest a structural hypothesis unifying these observations. Central to this model is the presumption of the existence of hydrogen bond-mediated interactions between metal ion-binding sites. 相似文献
2.
Salt-induced cooperativity in ATPase activity of plasma membrane-enriched fractions from cultured Citrus cells: kinetic evidence 总被引:1,自引:0,他引:1
ATPase activity was studied in plasma membrane-enriched fractions prepared from cultured Citrus sinensis L. cv. Osbeck cells. In general, properties of the plasma membrane ATPase from cultured cells, such as optimal pH and temperature. Vmax and Km were similar to those already observed in higher plants. The effects of high salt concentrations on ATPase activity were studied in membrane fractions derived from salt-sensitive and salt-tolerant cells grown in the presence or absence of salt. NaCl did not have an in vivo effect on Vmax and the apparent Km value for ATP. However, high concentrations of NaCl, or KCl, added in vitro, induced cooperativity in the enzyme and reduced the affinity of the enzyme for its substrate. Isoosmolar concentrations of sucrose or choline chloride failed to do so. Our results suggest that the plasma membrane ATPase of Citrus cells has more than one substrate-binding site on the native form of the enzyme which interact in the presence of salt and act independently in its absence. 相似文献
3.
The theory of interaction parameters has thus far been based on the free-energy relationships in the formation of ternary complexes formed between a pair of ligands and a protein molecule. The concept has been formulted in terms of a thermodynamic square comprised of the free protein, the two binary complexes, and the ternary complex. However, an increasing number of proteins have been found to exist as equilibrium mixtures of two macrostates. The equilibrium constants for such two-state transitions vary quite considerably between the various binary and ternary complexes of a given protein. We show here that the interpretations of interaction parameters in such two-state systems, requiring the use of a thermodynamic cube, are much more complex than those based on the classic thermodynamic square commonly employed. We demonstrate the use of enthalpies of interaction and heat capacities of interaction to analyze the source of observed free enerigies of interaction in such systems. Specifically, we find that measured negative interaction parameters may arise simply from the inability of a system to achieve all of the positive component effects anticipated by the conventional formulation. 相似文献
4.
Jacques Ricard Jacques Vergne Jean-Luc Decout Marie-Christine Maurell 《Journal of molecular evolution》1996,43(4):315-325
A polyallylamine carrying long hydrophobic dodecyl groups and adenine residues as side chains (PALAD C12) may be able to catalyze the hydrolysis ofN-carbobenzoxy-l-alaninep-nitrophenyl ester (N-Cbz-Ala) as well asp-nitrophenyl acetate (pNPA). The progress curve of hydrolysis of the former displays a long lag and apparently no steady state.
After this transient the rate falls off due to the accumulation of the products. Conversely, the hydrolysis ofp-nitrophenyl acetate displays classical burst kinetics followed by a slow decline of the reaction rate.
Theoretical considerations show that a steady state may be expected to occur only if the concentration of the free catalyst
is very small during the reaction. This condition is sufficient to allow the rate of disappearance of the substrate to be
equal to the rate of appearance of the products, which is precisely a condition for the existence of a steady state. If the
catalyst is poorly active and has a loose affinity for its substrate and product, the measurement of a significant reaction
rate will require a much larger concentration of the catalyst. Therefore, under these conditions, one cannot expect a steady
state to occur. The mathematical expression of the error made in the steady-state assumption has been derived. This error
increases with the catalyst concentration and decreases if the affinity of the substrate for the catalyst is high. Therefore
the lack of steady state is associated with the affinity (or the dissociation) of the substrate and the product for the catalyst.
When this affinity is low, the free concentration of the catalyst during the reaction is high and one cannot expect a steady
state to occur. This is precisely what takes place with N-Cbz-Ala.
A mathematical expression of the rate of hydrolysis of N-Cbz-Ala and of any reactant that displays this type of kinetics may
be derived at the end of the transient when the rate is close to its maximum value. Under these conditions the rate cannot
follow classical Michaelis-Menten kinetics and displays positive cooperativity.
It may therefore be speculated that primordial template-like catalysts that were displaying a poor affinity for their substrates
and products were already exhibiting apparent positive cooperativity in the kinetic reactions they were able to catalyze.
Correspondence to: J. Ricard 相似文献
5.
Stefan Mihailescu John M. Matsoukas Francis L. Abel Valeriu Nestianu Florin Romanescu Cristina Alecu-Parvu 《Letters in Peptide Science》1996,3(4):181-184
Summary The angiotensin II (ANG II) receptor blocker properties of sarmesin and its influence on the homotropic cooperativity of ANG II receptors were studied in two experimental models: isolated rabbit aorta and isolated rabbit atria. The results show that: (i) sarmesin is a specific competitive antagonist of ANG II receptors, with high affinity (pA2=8.93 in the isolated aorta and 8.66 in the isolated atria); and (ii) the slope of the concentration-response curves to ANG II and the Hill coefficient increased in the presence of sarmesin, the latter suggesting an enhancement of the positive homotropic cooperativity of ANG II receptors. These results may be explained overall by the reciprocal negative modulation of receptor affinity between sarmesin and ANG II, due, possibly, to opposite effects on the binding of G-proteins to ANG II receptors. 相似文献
6.
Minimal photosynthetic catalytic F1() core complexes, containing equimolar ratios of the and subunits, were isolated from membrane-bound spinach chloroplast CF1 and Rhodospirillum rubrum chromatophore RrF1. A CF1-33 hexamer and RrF1-11 dimer, which were purified from the respective F1() complexes, exhibit lower rates and different properties from their parent F1-ATPases. Most interesting is their complete resistance to inhibition by the general F1 inhibitor azide and the specific CF1 inhibitor tentoxin. These inhibitors were earlier reported to inhibit multisite, but not unisite, catalysis in all sensitive F1-ATPases and were therefore suggested to block catalytic site cooperativity. The absence of this typical property of all F1-ATPases in the 11 dimer is consistant with the view that the dimer contains only a single catalytic site. The 33 hexamer contains however all F1 catalytic sites. Therefore the observation that CF1-33 can bind tentoxin and is stimulated by it suggests that the F1 subunit, which is required for obtaining inhibition by tentoxin as well as azide, plays an important role in the cooperative interactions between the F1-catalytic sites.Abbreviations CF0F1
chloroplast F0F1
- CF1
chloroplast F1
- CF1
chloroplast F1 subunit
- CF1
chloroplast F1 subunit
- CF1()
a complex containing equal amounts of the CF1 and subunits
- MF1
mitochondrial F1
- RrF0F1
Rhodospirillum rubrum F0F1
- RrF1
R. rubrum F1
- RrF1
R. rubrum F1 subunit
- RrF1
R. rubrum F1 subunit
- RrF1()
a complex containing equal amounts of the RrF1 and subunits
- Rubisco
Ribulose-1,5-bisphosphate carboxylase
- TF1
thermophilic bacterium PS3 F1 相似文献
7.
K. A. Koehler M. K. Jain D. A. Gabriel H. -Y. C. Chang O. P. Malhotra 《Journal of Protein Chemistry》1995,14(7):537-548
The interaction of bovine prothrombin with Ca2+ and Mg2+ ions was investigated by following H+ release as a function of metal ion concentration at pH 6 and pH 7.4 at high and low ionic strength. Prothrombin Ca2+ and Mg2+ binding is characterized by high- and low-affinity sites. M2+ binding at these sites is associated with intramolecular conformational changes and also with intermolecular self-association. The pH dependence of H+ release by M2+ is bell shaped and consistent with controlling pKa values of 4.8 and 6.5. At pH 6 and low ionic strength, both Ca2+ and Mg2+ titrations following H+ release clearly show independent low- and high-affinity binding sites. Laser light scattering reveals that at pH 7.4 and low ionic strength, and at pH 6.0 and high ionic strength, the prothrombin molecular weight is between 73 and 98 kD. At pH 7.4 and high ionic strength, prothrombin is monomeric in the absence of metal ions, but appears to dimerize in the presence of M2+. At pH 6.0 and low ionic strength prothrombin exists as a dimer in the absence of metal ions and is tetrameric in the presence of Ca2+ and remains dimeric in the presence of Mg2+. These results and those for metal ion-dependent H+ release indicate that H+ release occurs concomitantly with association processes involving prothrombin.Abbreviations GLA
-carboxyglutamic acid; fragment 1. amino terminal residues 1–156 of bovine prothrombin
- MES
2-(N-morpholino) ethanesulfonic acid
- MOPS
3-(N-morpholino) propanesulfonic acid
- PS/PC
phosphatidylserine/phosphatidylcholine vesicles
-
ionic strength 相似文献
8.
S. W. Englander L. Mayne Y. Bai T. R. Sosnick 《Protein science : a publication of the Protein Society》1997,6(5):1101-1109
This discussion, prepared for the Protein Society's symposium honoring the 100th anniversary of Kaj Linderstrøm-Lang, shows how hydrogen exchange approaches initially conceived and implemented by Lang and his colleagues some 50 years ago are contributing to current progress in structural biology. Examples are chosen from the active protein folding field. Hydrogen exchange methods now make it possible to define the structure of protein folding intermediates in various contexts: as tenuous molten globule forms at equilibrium under destabilizing conditions, in kinetic intermediates that exist for less than one second, and as infinitesimally populated excited state forms under native conditions. More generally, similar methods now find broad application in studies of protein structure, energetics, and interactions. This article considers the rise of these capabilities from their inception at the Carlsberg Labs to their contemporary role as a significant tool of modern structural biology. 相似文献
9.
Alicia Alonso Colin J. Restall Margaret Turner Juan C. Gomez-Fernandez Felix M. Goñi Dennis Chapman 《生物化学与生物物理学报:生物膜》1982,689(2):283-289
Bacteriorhodopsin has been reconstituted at various molar concentrations into liposomes of dimyristoyl- and also of dipalmitoylphosphatidylcholine. Differential scanning calorimetry indicates that as the protein concentration within the lipid bilayer increases, the cooperativity of the lipid phase transition is reduced, i.e. the transition is broadened, while the midpoint transition temperature remains virtually unchanged. Freeze-fracture electron microscopy of our preparation shows, in agreement with previous data from other laboratories, that extensive protein aggregation occurs when the liposome is cooled below the Tc transition temperature of the lipid. Laser flash photolysis measurements of protein rotation of the bacteriorhodopsin show, especially in the case of protein-rich recombinants, that protein aggregates exist even above Tc. The perturbation caused by the presence of bacteriorhodopsin in the lipid bilayer is similar to that produced by other intrinsic proteins. The difficulty of correlating the observed calorimetric enthalpy data with a simple concept of a ‘boundary lipid layer’ based upon consideration of a single isolated protein is discussed in view of the occurrence of protein aggregates both above and below Tc. It is concluded that the reduction of enthalpy is related to the number of lipids which solvate the protein aggregates within the protein-lipid patches and are thereby removed from the cooperative melting and enthalpy of the remaining regions of pure lipid. 相似文献
10.
Paul Jensén 《Physiologia plantarum》1981,52(4):437-441
Uptake of Rb+ from a complete nutrient solution with 2.0 mM Rb+ was studied in roots of spring wheat seedlings ( Triticum aestivum L. cv. Svenno) with different K+ levels. The relationship between Rb+ uptake and concentration of K+ in the roots indicated a negative feedback mechanism operating through allosteric control. The Rb+ uptake process in root cells was divided into two steps: (1) binding of the ion in the free space, and (ii) transmembrane transport into the cytoplasm. Metabolic and non-metabolic components of uptake were separated by addition of the metabolic inhibitor 2,4-dinitrophenol (DNP) to the nutrient solution. It is suggested that metabolic Rb+ uptake requires energy in two uptake steps (for binding to the carrier entity in the free space and for transmembrane transport) or in one step only (for transmembrane transport), dependent on the K+ status of the roots. The change from metabolic to non-metabolic binding in the free space is accomplished by changing the conformational state of the carrier (slow/fast transitions). There may be a hysteretic effect on metabolic Rb+ uptake through a slow transition between carrier states. This is superimposed on the negative cooperativity, strengthening further cooperativity at intermediate K+ levels in the roots. Non-metabolic Rb+ uptake probably consists of two components, a carrier-mediated (facilitated diffusion) and a parallel diffusive component. 相似文献