PFG NMR studies of lysine-silica solutions

The formation process of silica nanoparticles in lysine-silica mixtures was studied using dynamic light scattering (DLS) and pulsed-field gradient (PFG) NMR measurements. (1)H NMR shows line broadening of the lysine resonances during TEOS hydrolysis/nanoparticle formation. Analysis of the TEOS hydrolysis kinetics show that TEOS hydrolysis is the rate-limiting step in particle formation, and has an activation energy of 20.5 kJ/mol. Transverse relaxation measurements show a corresponding decrease in T(2) with TEOS hydrolysis, indicating a reduction in the lysine mobility due to lysine-silica interactions. PFG NMR results indicate a systemic decrease in the self-diffusion coefficient of lysine as particle formation proceeds. The results obtained can be described using a two-state model wherein lysine is either free in solution or bound to the nanoparticles. Analysis of the PFG data of samples made at various temperatures show that lysine coverage upon complete hydrolysis is between 2.5 and 2.8 mmol lysine/kg solution, and insensitive to the heating temperature. PFG NMR shows a linear increase in the amount of bound lysine with increasing lysine content, indicating an increase in the surface area present, i.e. more and smaller particles, with increased lysine content. The PFG NMR results presented give quantitative insights that indicate that while pH is likely the primary driver for the rate of particle formation and particle size, lysine is critical for stabilization of the nanoparticles.     

赖氨酸胍基化对蛋白质组分析的影响

赖氨酸胍基化在蛋白质组学定性和定量研究中起着重要作用,本文系统分析了胍基化前后,HeLa细胞蛋白质经胰蛋白酶酶解产生的3种不同类型肽段的质谱鉴定情况,并探讨了不同肽段质谱响应改变的内在原因。发现赖氨酸在侧链能选择性地发生胍基化反应（其选择性达到96.8%）,转化为高精氨酸,碱性增强。因此在正离子质谱模式下,C端为赖氨酸的肽段产生了更多的y离子,提供了许多新的离子碎片信息。在鉴定结果中,此类肽段所占总肽段的比例由原来的51.7%上升为57.3%,并且有1015条新的肽段被检测到。对于不含有赖氨酸的肽段,其鉴定结果在胍基化前后基本没有变化。结果表明,胍基化可以在一定程度上提高质谱鉴定的灵敏度和互补性,提高蛋白质分析的覆盖率。     

Potentiometric sensor array for the determination of lysine in feed samples using multivariate calibration methods

A potentiometric sensor array has been developed for the determination of lysine in feed samples. The sensor array consists of a lysine biosensor and seven ion-selective electrodes for NH4+, K+, Na+, Ca2+, Mg2+, Li+, and H+, all based on all-solid-state technology. The potentiometric lysine biosensor comprises a lysine oxidase membrane assembled on an NH4+ electrode. Because the selectivity of the lysine biosensor towards other cation species is not sufficient, there is severe interference with the potentiometric response. This poor selectivity can be circumvented mathematically by analysis of the richer information contained in the multi-sensor data. The sensor array takes advantage of the cross-selectivity of lysine for each electrode, which differs from the other species and quantification of lysine in complex feed sample extracts is accomplished with multivariate calibration methods, such as partial least-squares regression. The results obtained are in a reasonable agreement with those given by the standard method for amino acid analysis.     

Available lysine in foods: a brief historical overview

A brief historical overview is presented on the development of the science addressing lysine bioavailability in foods. Early observations that dietary protein utilization did not always correlate with gross amino acid composition led to an understanding that the amino acid lysine, in particular, can be easily damaged during food processing. Conventional amino acid analysis, involving a strong acid hydrolysis step, can lead to a significant degree of overestimation of lysine in processed foods. More recently, it has been found that not only food lysine content values but also estimates of lysine digestibility and digestible lysine contents may be erroneous. Estimates of absorbed (true ileal digestible) reactive lysine are accurate measures of available lysine. Technically, bioassays such as the slope-ratio assay determine utilized rather than available lysine.     

ATR-IR spectroscopic study of L-lysine adsorption on amorphous silica

Attenuated total reflectance infrared (ATR-IR) spectroscopy was employed to quantitatively evaluate the dissociation states (di-cationic, cationic, zwitterionic, and anionic) of lysine adsorbed on amorphous silica. To determine the relationship between the ATR-IR spectra and each dissociation state, we first measured pH-induced spectral changes of dissolved lysine and correlated these changes with the thermodynamically calculated dissociation states of lysine. This procedure yielded calibration curves with good linearity; we used these curves for the quantitative analysis of adsorbed lysine. Our analysis revealed that 81+/-5% of the lysine adsorbed on amorphous silica was present in a cationic state and 19+/-5% was in a zwitterionic state; these percentages remained mostly unchanged over the whole range of pH values tested (pH = 7.1-9.8). We interpret the values obtained to indicate that lysine adsorption is mainly driven by electrostatic interaction with the negatively charged silica surface (SiO(-)...Lys(+), SiO(-)...Lys(+/-)).     

银二氧化硅核壳增强异硫氰酸荧光素酯滤纸基质室温磷光法检测赖氨酸

银二氧化硅核壳( Ag@ SiO2)可使异硫氰酸荧光素酯(FITC)产生金属增强室温磷光(MERTP).当所研究体系加入赖氨酸时,其MERTP猝灭.据此,建立了检测赖氨酸的新方法.在FITC和Ag@ Si02的加入量之比为1∶1.5,pH- 8.0时,本方法检测赖氨酸的检出限为0.270 μmol/L;线性范围为1.50～90.5 μmol/L,人二合成样品加标回收率为97.0％～108.0％;相对标准偏差为2.1％(n=9).本方法用于检测赖氨葡锌颗粒和赖氨酸磷酸氢钙片中的赖氨酸,结果满意.     

Interaction of guanine, its anions, and radicals with lysine in different charge states

Modification in DNA or protein structure can severely affect DNA-protein interactions and the functioning of biological systems. Some new insights into radiation-induced effects of guanine-lysine interactions have been obtained here by theoretical investigations. Geometries of zwitterionic and non-zwitterionic lysine in different charge states (neutral, radical cation, and protonated cation) were optimized employing the B3LYP/6-31G** and B3LYP/AUG-cc-pVDZ levels of hybrid density functional theory (DFT) and using the second-order M?ller-Plesset perturbation theory along with the 6-31G** basis set. In the case of neutral lysine in the gas phase, no zwitterionic structure was obtained. The non-zwitterionic structures of lysine in radical and protonated cationic forms are appreciably more stable than the corresponding zwitterionic structures in the gas phase as obtained at all levels of theory employed here. Binding of guanine and different dehydrogenated guanine radicals with lysine in different charge states was studied at the B3LYP/6-31G** level of DFT. When guanine makes a complex with the lysine radical cation, large amounts of spin and positive charge densities are transferred from the lysine radical cation to guanine and the guanine is thus converted from its normal form to the radical cationic form. Complexation of the lysine radical cation with the H1-hydrogen-abstracted guanine radical leads to CO2 liberation and proton transfer from lysine. These results are compared with the available experimental ones.     

Oligo-lysine Induced Formation of Silica Particles in Neutral Silicate Solution

Introduction Theformationofsilicastructuresunderthecontrol ofalivingorganism(biosilification)isawidespread phenomenoninnature,whichisfoundinmanylife formssuchasdiatoms,spongesandgrasses,etc.[1,2].Themostattractiveaspectofbiosilificationisthatex quisitestr…     

A comparison of the mechanism for the reductive half-reaction between pea seedling and other copper amine oxidases (CAOs)

In a previous DFT study a mechanism for the reductive half-reaction of pea seedling amine oxidase (PSAO) was suggested. In many of the suggested steps a lysine at the active site plays an important role. However, this lysine is not found in other amine oxidases. The primary aim of the present DFT study is therefore to investigate alternative mechanisms for those amine oxidases (CAO) where the lysine residue is not present. One of the most important roles suggested for the lysine in PSAO was to protonate the O2-site of TPQ before the critical Cbond;H bond cleavage of the substrate. In the absence of lysine the O2-site of TPQ is now suggested to be protonated by a water ligand on the copper metal complex, in line with experimental suggestions. In other steps the role of lysine is taken over by an asparagine. All results are compared with experimental observations and good agreement is generally found.     

The uncertainty in the calculation of the amount of blocked and reactive lysine in milk products,as determined by the furosine method

The measurement uncertainty in the calculation of the amount of blocked and reactive lysine (as determined by the furosine method) was evaluated according to the procedure described in the Eurachem/CITAC guide. The analytical method involves the chromatographic determination of lysine and furosine after acid hydrolysis. The calculation of blocked and reactive lysine in the initial protein is based on known conversion factors. The estimation of the uncertainty was performed in two steps: (1) determination of the uncertainty in the chromatographic determination of lysine and furosine, and (2) determination of the uncertainty in the calculation of blocked and reactive lysine. The individual contributions to the final uncertainty were identified, quantified, and combined in uncertainty budgets. The largest contribution to the calculation of blocked lysine came from estimating the conversion factor of blocked lysine into furosine during acid hydrolysis. For the calculation of reactive lysine, the main contribution came from the chromatographic determination of lysine. The uncertainty estimates were compared to available validation data (in-house and collaborative standard deviations of reproducibility).     

Flow injection differential potentiometric determination of lysine by using a lysine biosensor

This paper describes a method for the determination of lysine based on a flow injection (FI) differential potentiometry system. The flow injection manifold is composed of a support electrolyte solution channel and a water channel acting as a carrier into which the sample solution is injected. The lysine biosensor was consisted of lysine oxidase chemically immobilized on a nylon membrane and attached to an all-solid-state ammonium electrode. A circular ammonium electrode was used as a reference. Hence, the possible interference of endogenous ammonium can be partly corrected by differential potentiometry. In order to increase the sensitivity of the response, the reaction was kinetically developed following a stopped-flow method. As a result, the sensitivity increased from 20 to 40 mV per decade when comparing the FI and the stopped-flow values. Furthermore, the peak-to-peak stopped-flow signals generated can be used as a more selective analytical response for lysine. The quantification of lysine in mixture samples containing small amounts of ammonium can be achieved with an acceptable accuracy, with prediction errors lower than 4%. However, when the ammonium concentration exceeded the lysine concentration, multivariate calibration with non-linear partial least squares (PLS) regression was needed to improve the lysine quantification, with an overall prediction error around 10%.     

赖氨酸的毛细管电泳特性及测定

研究了缓冲液pH和操作温度对赖氨酸毛细管区带电泳分离中运行电流、电渗迁移率、峰形等参数的影响,确定了赖氨酸电泳分离的最佳实验条件,建立了毛细管区带电泳测定赖氨酸的方法。线性范围为0．1－3．0g/L,检出限为0．03g/L。该法操作简便迅速,重复性好。用于赖氨酸发酵液中赖氨酸的含量测定,结果令人满意。     

Electrospray ionisation tandem mass spectrometric study of hydrogen-bonding interactions of some disaccharides with lysine

The efficiency of formation of protonated heterotrimers of lysine with underivatised sugars (mono-, di-, and trisaccharides) and N-acetylglucosamine (N-AcGlc) was studied under electrospray ionisation conditions. The collision induced dissociation spectra of [Lys + sugar + NAcGlc + H](+) resulted in [Lys + NAcGlc + H](+) and [Lys + sugar + H](+) as the major product ions. Relative abundances of these two fragments reflect the extent of adduct formation of protonated lysine plus sugar, with reference to the reference compound NAcGlc. This relative abundance ratio was found to be characteristic of the sugar structure. In this way it was observed that the ability of lysine to form a protonated heterodimer with neutral sugars increases with an increase in the number of acetal oxygens. Lactose showed an anomalously high affinity for protonated Lys, possibly reflecting the axial hydroxyl at C4. The postulated involvement of the epsilon-NH(2) group of lysine in the formation of protonated heterodimers with sugars was supported by similar results of similar experiments with NH(3) in place of lysine.     

Helical peptide models for protein glycation: proximity effects in catalysis of the Amadori rearrangement.

INTRODUCTION: Non-enzymatic glycation of proteins has been implicated in various diabetic complications and age-related disorders. Proteins undergo glycation at the N-terminus or at the epsilon-amino group of lysine residues. The observation that only a fraction of all lysine residues undergo glycation indicates the role of the immediate chemical environment in the glycation reaction. Here we have constructed helical peptide models, which juxtapose lysine with potentially catalytic residues in order to probe their roles in the individual steps of the glycation reaction. RESULTS: The peptides investigated in this study are constrained to adopt helical conformations allowing residues in the i and i+4 positions to come into spatial proximity, while residues i and i+2 are far apart. The placing of aspartic acid and histidine residues at interacting positions with lysine modulates the steps involved in early peptide glycation (reversible Schiff base formation and its subsequent irreversible conversion to a ketoamine product, the Amadori rearrangement). Proximal positioning of aspartic acid or histidine with respect to the reactive lysine residue retards initial Schiff base formation. On the contrary, aspartic acid promotes catalysis of the Amadori rearrangement. Presence of the strongly basic residue arginine proximate to lysine favorably affects the pK(a) of both the lysine epsilon-amino group and the singly glycated lysine, aiding in the formation of doubly glycated species. The Amadori product also formed carboxymethyl lysine, an advanced glycation endproduct (AGE), in a time-dependent manner. CONCLUSIONS: Stereochemically defined peptide scaffolds are convenient tools for studying near neighbor effects on the reactivity of functional amino acid sidechains. The present study utilizes stereochemically defined peptide helices to effectively demonstrate that aspartic acid is an efficient catalytic residue in the Amadori arrangement. The results emphasize the structural determinants of Schiff base and Amadori product formation in the final accumulation of glycated peptides.     

Preparation and evaluation of a lysine-bonded silica monolith as polar stationary phase for hydrophilic interaction pressurized capillary electrochromatography

A silica-based monolith as polar stationary phase was described for hydrophilic interaction pressurized capillary electrochromatography (HI-pCEC). The polar monolithic column was prepared by on-column reaction of lysine with epoxy groups on a gamma-glycidoxypropyltrimethosysilane-modified silica monolith. The stationary phase yielded strong hydrophilic interaction due to the slightly polar hydroxyl groups, and the strong polar lysine ligand with amino groups and carboxylic groups contained on the surface of the monolith. In order to evaluate the hydrophilic character of lysine ligand, the chromatographic behaviors of epoxy monolith (before lysine bonded) and diol monolith (hydroxyl groups contained) were also investigated. Two groups of comparative experiment were developed in terms of the separation of typical neutral non-polar and polar compounds performed in a mobile phase of aqueous-acetonitrile solution. Results showed that the lysine monolith was much more hydrophilic than the diol monolith, which presented less hydrophobic than the epoxy monolith. For further study on its hydrophilic character, the lysine monolith was demonstrated in the HI-pCEC mode for the separations of various polar compounds such as phenols, nucleic acid bases and nucleosides.     

赖氨酸与香草醛反应产物的极谱波特征及其分析应用

研究了赖氨酸与香草醛反应产物的极谱波特征并建立了赖氨酸的分析方法。在0.1mol/L磷酸盐缓冲溶液中赖氨酸与香草醛反应的产物于－1.16V产生灵敏的极谱波,赖氨酸浓度在5×10     

In situ ATR-IR investigation of L-lysine adsorption on montmorillonite

The adsorption behavior of lysine on montmorillonite in aqueous solution was investigated by in situ attenuated total reflectance infrared (ATR-IR) spectroscopy. To distinguish the protonation states of α-amino group, side-chain amino group and carboxyl group in lysine structure using ATR-IR spectra (i.e., NH₂ versus and COO⁻ versus COOH), pH-induced spectral changes of dissolved lysine were firstly measured and correlated with the thermodynamically calculated dissociation states of lysine (di-cationic, cationic, zwitterionic and anionic states). The obtained result was applied to interpret the ATR-IR spectra of lysine adsorbed on montmorillonite. We found that the adsorbed lysine was dominantly present as cationic state over the whole range of tested pH (pH = 4.9–9.7). This indicates that the adsorption is mainly driven by electrostatic interaction between the negatively charged montmorillonite surface and positively charged cationic lysine. We also found that lysine interacts with montmorillonite surface through the protonated side-chain amino group. This result suggests that lysine has a preferred vertical orientation, with the side-chain amino group pointing toward the surface.     

para-Hydrogenation of unsaturated moieties on poly(lysine) derived substrates for the development of novel hyperpolarized MRI contrast agents

Four alkyne-functionalized poly(lysine) derivatives have been synthesized and characterized by 1H and 13C NMR spectroscopy. In the first poly(lysine) derivative, phenylpropiolate moieties are directly bound to the aminic arms, whereas in the other derivatives, propargylamine moieties are bound to the aminic poly(lysine) arms through glucaric acid and diethylene glycol (DG) chains, respectively. para-Hydrogenation of the alkyne-functionalized poly(lysine) compounds has been investigated and the results have been discussed in terms of spin lattice relaxation properties of the hydrogenated products. It is shown that the longer the mobile chain separating the unsaturation from the poly(lysine) backbone, the more intense the polarized signals when para-hydrogenation is carried out. This is due to (a) the maintenance of short reorientational times on the unsaturated ends, and therefore a sufficiently long T(1) of the protons added during hydrogenation, and (b) the minor effect of steric hindrance by the poly(lysine) backbone that decreases interaction of the unsaturation with the catalyst, allowing higher hydrogenation rates.