scholarly journals Substitution of lanthanide ions for calcium ions in the activation of bovine prothrombin by activated factor X. High affinity metal-binding sites of prothrombin and the derivatives of prothrombin activation.

1976 ◽  
Vol 251 (11) ◽  
pp. 3235-3241
Author(s):  
B C Furie ◽  
K G Mann ◽  
B Furie
Keyword(s):  
Calcium Ions ◽  
Metal Binding ◽  
Binding Sites ◽  
Lanthanide Ions ◽  
Factor X ◽  
Metal Binding Sites ◽  
High Affinity ◽  
Derivatives Of ◽  
Prothrombin Activation

DETERMINATION OF NUMBER OF γ-CARBOXYGLUTAMIC ACID (GLA) RESIDUES INVOLVED IN FORMING THE TWO HIGH AFFINITY METAL BINDING SITES IN PROTHROMBIN AND FACTOR X

1987 ◽  
Author(s):  
G L Brodsky ◽  
S P Bajaj
Keyword(s):  
Metal Binding ◽  
Binding Sites ◽  
Ion Binding ◽  
Factor X ◽  
Metal Binding Sites ◽  
High Affinity ◽  
Carboxyglutamic Acid ◽  
High Affinity Site ◽  
Ion Binding Sites

Prothrombin and factor X possess two high affinity and several low affinity lanthanide ion binding sites. In both proteins, the association constant of the high affinity sites is at least 50-fold greater than that of the low affinity sites. Moreover, metal bound to these high affinity sites is extremely difficult to displace. It has been proposed that one of the two high affinity sites in factor X involves Gla residues while the other involves β-hydroxyaspartic acid and no Gla residues. It is also known that ^H can be specifically incorporated into Gla residues at an acidic pH. We have determined that under nondenaturing conditions when Gla (synthetic or in proteins) is complexed to metal at pH 5.5, this specific 3H incorporation is blocked. Furthermore, we have found that β-hydroxyaspartic acid does not incorporate in the presence or absence of metal. When we incubated prothrombin or factor X (41 μM) with 3H2O in the presence of Tb3+ or Gd3+ (82 μM), we blocked 5.6 Gla residues per prothrombin and 5.5 Gla residues per factor X from 3H incorporation. Under these conditions, we calculated that >95% of the high affinity sites are occupied by metal. Thus, in prothrombin, an average of 2.8 Gla residues are involved in forming each high affinity site. If the Gla residues in factor X participate in forming only one of the two high affinity sites, then all 5.5 Gla residues blocked from incorporation must be involved in forming that site. However, this seems highly unlikely. We conclude that, as in prothrombin, both high affinity sites in factor X involve Gla residues (average 2.75/site). However, our data does not exclude the possibility of existence of a heterologous site containing both β-hydroxyaspartic acid and Gla residues.

scholarly journals Effects of Multiple Metal Binding Sites on Calcium and Magnesium-dependent Activation of BK Channels

2005 ◽  
Vol 127 (1) ◽  
pp. 35-50 ◽  
Author(s):  
Lei Hu ◽  
Huanghe Yang ◽  
Jingyi Shi ◽  
Jianmin Cui
Keyword(s):  
Metal Binding ◽  
Binding Sites ◽  
Quantitative Estimation ◽  
Bk Channels ◽  
Channel Activation ◽  
Metal Binding Sites ◽  
High Affinity ◽  
A Site ◽  
Metal Sensing ◽  
Allosteric Model

BK channels are activated by physiological concentrations of intracellular Ca2+ and Mg2+ in a variety of cells. Previous studies have identified two sites important for high-affinity Ca2+ sensing between [Ca2+]i of 0.1–100 μM and a site important for Mg2+ sensing between [Mg2+]i of 0.1–10 mM. BK channels can be also activated by Ca2+ and Mg2+ at concentrations >10 mM so that the steady-state conductance and voltage (G-V) relation continuously shifts to more negative voltage ranges when [Mg2+]i increases from 0.1–100 mM. We demonstrate that a novel site is responsible for metal sensing at concentrations ≥10 mM, and all four sites affect channel activation independently. As a result, the contributions of these sites to channel activation are complex, depending on the combination of Ca2+ and Mg2+ concentrations. Here we examined the effects of each of these sites on Ca2+ and Mg2+-dependent activation and the data are consistent with the suggestion that these sites are responsible for metal binding. We provide an allosteric model for quantitative estimation of the contributions that each of these putative binding sites makes to channel activation at any [Ca2+]i and [Mg2+]i.

Metal Binding to Pseudomonas aeruginosa Azurin: a Kinetic Investigation

2000 ◽  
Vol 55 (5-6) ◽  
pp. 347-354 ◽  
Author(s):  
Fabio Naro ◽  
Maria G. Tordi ◽  
Giorgio M. Giacometti ◽  
Francesco Tomei ◽  
Anna M. Timperio ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Metal Binding ◽  
Binding Sites ◽  
Fluorescence Emission ◽  
Uv Spectrum ◽  
Metal Binding Site ◽  
Metal Binding Sites ◽  
High Affinity ◽  
Redox Center ◽  
Sh Group

The interaction between azurin from Pseudomonas aeruginosa and Ag(I), Cu(II), Hg(II), was investigated as a function of protein state, i.e. apo-, reduced and oxidised azurin. Two different metal binding sites, characterized by two different spectroscopic absorbancies, were detected: one is accessible to Ag(I) and Cu(II) but not to Hg(II); the other one binds Ag(I) and Hg(II) but not copper. When added in stoichiometric amount, Ag(I) shows high affinity for the redox center of apo-azurin, to which it probably binds by the -SH group of Cys112; it can displace Cu(I) from reducedazurin, while it does not bind to the redox center of oxidizedazurin. Kinetic experiments show that Ag(I) binding to the reducedform is four times faster than binding to the apo-form. This result suggests that metal binding requires a conformational rearrangement of the active site of the azurin. Interaction of A g(I) or Hg(II) ions to the second metal binding site, induces typical changes of UV spectrum and quenching of fluorescence emission.

The Binding of Calcium Ions to Bovine Factor X by Rate Dialysis

1978 ◽  
Vol 40 (02) ◽  
pp. 350-357
Author(s):  
Robert H Yue ◽  
Menard M Gertler
Keyword(s):  
Molecular Weight ◽  
Dissociation Constant ◽  
Calcium Ions ◽  
Activation Process ◽  
Factor X ◽  
The Real ◽  
High Affinity ◽  
Binding Data ◽  
Sequential Mechanism ◽  
Bovine Factor

SummaryThe binding of Ca+2 to bovine factor X (molecular weight of 74,000) (Yue und Gertler 1977) was studied by the technique of rate dialysis and with the use of 45Ca+2. The binding data are consistent with a model of sequential mechanism. One mole of Ca+2 binds to the glycoprotein with a dissociation constant of 5.2 × 10-5 M and an additional 39 ± 4 moles of Ca+2 bind to this zymogen with a dissociation constant of 3.7 × 10-3M. The binding of the high affinity Ca+2 causes a functionally significant change in the zymogen, and (calcium) (factor X) complex is the real substrate in the activation process by the protease in Russell’s viper venom.

scholarly journals Rational design of metal-binding sites in domain-swapped myoglobin dimers

2021 ◽  
Vol 217 ◽  
pp. 111374
Author(s):  
Satoshi Nagao ◽  
Ayaka Idomoto ◽  
Naoki Shibata ◽  
Yoshiki Higuchi ◽  
Shun Hirota
Keyword(s):  
Metal Binding ◽  
Binding Sites ◽  
Rational Design ◽  
Metal Binding Sites
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