Dynamic ionic radius of alkali metal ions in aqueous solution: a pulsed-field gradient NMR study

Stokes radius (dynamic ionic radius) of the alkali metal ions versus the ionic radius (Rion) at 303 K. The dotted line is a guide for the 1 : 1 relation.

Hydration Thermodynamics and Hydrodynamic (Stokes) Radius of the Lanthanide Ions

Many biochemical and physiological properties depend on the size of ions and the thermodynamic quantities of ion hydration. The diffusion coefficient (D) of lanthanide (III) ions (Ln+3) in solution assumed (1.558-1.618 ×10−9 m2 s−1) by Einstein–Smoluchowski relation. The association constant (KA) of Ln+3 ions was calculated (210.3-215.3 dm3 mole-1) using the Shedlovsky method, and the hydrodynamic radius calculated (1.515-1.569 ×10−10 m) by the Stokes-Einstein equation. The thermodynamic parameters (ΔGo, ΔSo) also calculated by used suitable relations, while ΔHo, values are obtained from the literature. ΔGo, for ion hydration, has negative values in the range (13.25-13.30 KJ/mole), and a negative ΔSo, results have been shown in the limit (11.016-12.506 KJ/ K. mole).

P1100IMPACT OF THE RELATIONSHIP BETWEEN LOW MOLECULAR WEIGHT PROTEINS REMOVAL EFFICIENCIES AND THE VOLUME OF ALBUMIN LEAKAGE IN HEMODIAFILTRATION THERAPY

Background and Aims The purpose of hemodiafiltration (HDF) is to remove small- to large molecular weight solutes in order to prevent and treat complications and to improve the prognosis of dialysis patients. Especially, larger low molecular weight proteins (LMWPs) must be removed efficiently. β2-microglobulin (MG) (molecular weight [MW], 11.8 kDa: Stokes radius, 15.6A), is considered as an important marker of dialysis efficiency. However, MW of β2-MG is too small as a marker in HDF. Conversely, α1-MG (MW, 33 kDa: Stokes radius, 28.4A) is the most appropriate marker for evaluation of solute removal efficiency of HDF because of the following reasons: (1) optical molecular size; (2) not very low physiological blood concentration; (3) stable production rate; (4) removal by convection; and (5) accumulation in ESRD. Efficient removal of α1-MG without losing albumin (Alb) (MW,66 kDa: Stokes radius, 35.5A) is impossible even by HDF. In this study, we investigated the relationship between LMWP removal efficiencies and the volume of Alb leakage by analyzing the results of patients undergoing HDF. And we also evaluated the trend of the serum levels of patients who had received high-efficiency HDF for more than three years Method Eighty-seven patients who had received HDF in 2018–2019 were included. The results of HDF (a total of 435 sessions) were analyzed (blood flow rate, 261.5±28.5 mL/min; replacement fluid volume, 48.7 L/s pre-dilution on-line HDF and 14.2 L/s post-dilution on-line HDF). The removal efficiencies of β2-MG, prolactin (PRL), and α1-MG, and the volume of Alb leakage were examined. In addition, the relationship between the removal efficiency of each solute and the volume of Alb leakage was investigated. Changes in serum Alb level were analyzed in 13 patients who continued to receive high-efficiency HDF (the removal rate of α1-MG was 30% or more) for more than three years. Results Analysis of the results (mean ± SD) obtained from a total of 435 sessions (87 patients) showed that the removal rates of β2-MG, PRL, and α1-MG were 80.7 ± 4.5%, 75.8±9.4 and 33.8 ± 9.4%, respectively, and the Alb leakage was 3.9 ± 1.8 g/s. The equations of the approximate curve between Alb leakage and β2-MG, PRL, and α1-MG removal rates were expressed as follows: ï½™= 1.4197ln(x) + 78.972 (R2 = 0.0426), y = 9.7185ln(x) + 64.134 (R2 =0.4632), and ï½™™= 11.225ln(x) + 20.282 (R2 = 0.6191), respectively. These results indicate that the dynamics between Alb leakage and α1-MG removal are very similar. That is, although the molecular weight of Alb is twice as that of α1-MG, the difference in their Stokes radius is 20%; therefore, separate removal of Alb and α1-MG is impossible. The mean serum Alb level in 13 patients who received high-efficiency HDF for 3 years or longer was 3.59 ± 0.23 g/dL in January 2017 and 3.69 ± 0.28 g/dL in December 2019, revealing no changes. Conclusion The efficient removal of larger LMWPs leads to the leakage of Alb. However, an Alb leakage of 3 to 6 g/s has little influence on the serum Alb level of patients. Thus, α1-MG is the most appropriate marker of the solute removal efficiency of HDF.

Glucose deprivation activates a cAMP-independent protein kinase from Trypanosoma equiperdum

Kemptide (sequence: LRRASLG) is a synthetic peptide holding the consensus recognition site for the catalytic subunit of the cAMP-dependent protein kinase (PKA). cAMP-independent protein kinases that phosphorylate kemptide were stimulated in Trypanosoma equiperdum following glucose deprivation. An enriched kemptide kinase-containing fraction was isolated from glucose-starved parasites using sedimentation throughout a sucrose gradient, followed by sequential chromatography on diethylaminoethyl-Sepharose and Sephacryl S-300. The trypanosome protein possesses a molecular mass of 39.07–51.73 kDa, a Stokes radius of 27.4 Ǻ, a sedimentation coefficient of 4.06 S and a globular shape with a frictional ratio f/fo = 1.22–1.25. Optimal enzymatic activity was achieved at 37 °C and pH 8.0, and kinetic studies showed Km values for ATP and kemptide of 11.8 ± 4.1 and 24.7 ± 3.8 µm, respectively. The parasite enzyme uses ATP and Mg2+ and was inhibited by other nucleotides and/or analogues of ATP, such as cAMP, AMP, ADP, GMP, GDP, GTP, CTP, β,γ-imidoadenosine 5′-triphosphate and 5′-[p-(fluorosulfonyl)benzoyl] adenosine, and by other divalent cations, such as Zn2+, Mn2+, Co2+, Cu2+, Ca2+ and Fe2+. Additionally, the trypanosome kinase was inhibited by the PKA-specific heat-stable peptide inhibitor PKI-α. This study is the first biochemical and enzymatic characterization of a protein kinase from T. equiperdum.

The conformational state of polyphenol oxidase from field bean (Dolichos lablab) upon SDS and acid-pH activation

Field bean (Dolichos lablab) contains a single isoform of PPO (polyphenol oxidase) – a type III copper protein that catalyses the o-hydroxylation of monophenols and oxidation of o-diphenols using molecular oxygen – and is a homotetramer with a molecular mass of 120 kDa. The enzyme is activated manyfold either in the presence of the anionic detergent SDS below its critical micellar concentration or on exposure to acid-pH. The enhancement of kcat upon activation is accompanied by a marked shift in the pH optimum for the oxidation of t-butyl catechol from 4.5 to 6.0, an increased sensitivity to tropolone, altered susceptibility to proteolytic degradation and decreased thermostability. The Stokes radius of the native enzyme is found to increase from 49.1±2 to 75.9±0.6 Å (1 Å=0.1 nm). The activation by SDS and acid-pH results in a localized conformational change that is anchored around the catalytic site of PPO that alters the microenvironment of an essential glutamic residue. Chemical modification of field bean and sweet potato PPO with 1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide followed by kinetic analysis leads to the conclusion that both the enzymes possess a core carboxylate essential to activity. This enhanced catalytic efficiency of PPO, considered as an inducible defence oxidative enzyme, is vital to the physiological defence strategy adapted by plants to insect herbivory and pathogen attack.

The ABC transporter BmrA from Bacillus subtilis is a functional dimer when in a detergent-solubilized state

BmrA from Bacillus subtilis is a half-size ABC (ATP-binding cassette) transporter involved in multidrug resistance. Although its supramolecular organization has been investigated after reconstitution in a lipid bilayer environment, and shows a dimeric and possibly a tetrameric form, the precise quaternary structure in a detergent-solubilized state has never been addressed. In the present study, BmrA was purified from Escherichia coli membranes using an optimized purification protocol and different detergents. Furthermore, the ATPase activity of BmrA and the quantity of bound lipids and detergent were determined, and the oligomeric state was analysed using SEC (size-exclusion chromatography) and analytical ultracentrifugation. The activity and the quaternary structure of BmrA appeared to be strongly influenced by the type and concentration of the detergent used. SEC data showed that BmrA could be purified in a functional form in 0.05 and 0.01% DDM (n-dodecyl-β-D-maltoside) and was homogeneous and monodisperse with an Rs (Stokes radius) of 5.6 nm that is compatible with a dimer structure. Sedimentation-velocity and equilibrium experiments unequivocally supported that BmrA purified in DDM is a dimer and excluded the presence of other oligomeric states. These observations, which are discussed in relation to results obtained in proteoliposomes, also constitute an important first step towards crystallographic studies of BmrA structure.