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At least 235 records · Page 13Linked to original sources
PubMed1997

Binding kinetics of Bacillus sphaericus binary toxin to midgut brush-border membranes of Anopheles and Culex sp. mosquito larvae.

Direct-binding assays and homologous-competition assays were used to identify specific binding between the radiolabelled toxin of Bacillus sphaericus and brush-border membrane fractions (BBMF) from Anopheles gambiae and Anopheles stephensi, obtained from whole larvae preparations. In both species, the toxin bound to a single class of receptors. BBMF of A. gambiae had the highest binding affinity for the toxin of the species tested, with a dissociation constant (Kd) of 30 +/- 15 nM and a maximum receptor concentration of 5 +/- 1 pmol/mg. Toxin binding to A. gambiae BBMF was compared with that to BBMF from B. sphaericus-susceptible (IP) and B. sphaericus-resistant (SPHAE) Culex pipiens populations. BBMF toxin binding was slower in A. gambiae than in the C. pipiens populations. The BBMF of the B. sphaericus-resistant population of C. pipiens had an association profile that was similar to the susceptible population, despite of the lack of susceptibility in vivo. No relationship between toxicity and irreversibility of toxin binding was detected. On the contrary, toxin dissociation from BBMF was fast and almost complete in BBMF of all species studied.

Animals↗
PubMed1976

Tin-protein binding kinetics in normal and uremic plasma and its effect on dialysis fluxes.

A high affinity of plasma protein for tin has been demonstrated by ultrafiltration studies. At physiologic concentrations, UF/P remained below 1%, consistent with more than 99% binding. Binding of tin was nearly complete despite a decrease in plasma protein concentration to 3.5 Gm% and an increase in plasma tin by 80 mg/L. This binding inhibits efflux of tin from plasma during dialysis while influx occurs readily. This may be a mechanism for accumulation of tin in patients undergoing maintenance hemodialysis. A mechanism for tin chelation could be clinically beneficial in the syndrome of dialysis dementia.

Blood Proteins↗
PubMed1996

A fractal analysis of the influence of non-specific binding on antigen-antibody binding kinetics for biosensor applications.

A fractal analysis of the influence of non-specific binding on the specific binding of antigen in solution to antibody immobilized on a biosensor surface is presented for first-, one and a half-, second, and other-order reactions occurring under external diffusion-limited conditions. Both single-step and dual-step binding of antigen in solution to antibody immobilized on the surface is considered. For a first-order reaction, an increase in the fractal parameters, b, leads to a decrease in the amount of antigen in solution bound specifically to the antibody on the surface when non-specific binding is either absent or present. The presence of non-specific binding leads to a decrease in the amount of antigen bound to the antibody on the surface. For a one and a half- and for second-order reactions and when non-specific binding is either absent or present to a small degree (alpha = 0.01), an increase in the fractal parameter, b, leads to a decrease in the amount of antigen bound specifically to the antibody immobilized on the biosensor surface. However, for an alpha value of 0.1, the maximum rate and the amount of antigen bound specifically to the antibody immobilized on the biosensor surface is obtained for fractal parameter values of 0.2 and 0.4, and 0.4 for the one and a half- and for second-order reactions, respectively. Apparently, some amount of heterogeneity is helpful in obtaining the optimum amount and rate of antigen in solution bound specifically to the antibody on the surface for reaction orders higher than one. The applicability of the approach to real antibody surfaces is demonstrated.

Antigen-Antibody Reactions↗
PubMed2004

Anthrax toxin protective antigen: inhibition of channel function by chloroquine and related compounds and study of binding kinetics using the current noise analysis.

Protective antigen (PA) of the tripartite anthrax toxin binds to a cell surface receptor and mediates the transport of two enzymatic components, edema factor and lethal factor, into the cytosol of host cells. Here recombinant PA(63) from Bacillus anthracis was reconstituted into artificial lipid bilayer membranes and formed ion permeable channels. The heptameric PA(63)-channel contains a binding site for 4-aminoquinolones, which block ion transport through PA in vitro. This result allowed a detailed investigation of ligand binding and the stability constants for the binding of chloroquine, fluphenazine, and quinacrine to the binding site inside the PA(63)-channel were determined using titration experiments. Open PA(63)-channels exhibit 1/f noise in the frequency range between 1 and 100 Hz, whereas the spectral density of the ligand-induced current noise was of Lorentzian type. The analysis of the power density spectra allowed the evaluation of the on- and off-rate constants (k(1) and k(-1)) of ligand binding. The on-rate constants of ligand binding were between 10(6) and 10(8) M(-1) s(-1) and were dependent on the ionic strength of the aqueous phase, sidedness of ligand addition, as well as the orientation and intensity of the applied electric field. The off-rates varied between approximately 10 s(-1) and 2600 s(-1) and depended mainly on the structure of the ligand.

Algorithms↗
PubMed1995

Substrate and product binding sites of yeast fatty acid synthase. Stoichiometry and binding kinetics of wild-type and in vitro mutated enzymes.

The four known substrate binding sites of yeast fatty acid synthase (FAS), Ser819 (acetyltransferase, OHAC) and Ser5421 (malonyl/palmitoyl transferase, OHMa1) of subunit beta and Ser180 (pantetheine binding site, SHc) and Cys1305 (3-oxoacyl synthase, SHp) of subunit alpha were replaced, by targeted in vitro mutagenesis, by the non-acylatable amino acids glutamine, glycine or alanine. The four mutated FAS proteins together with two pairs of double mutants (OHAc/OHMa1 and SHc/SHp) were episomally expressed in appropriate delta fas1 or delta fas2 deletion strains. The purified enzymes isolated from these transformants were used for comparative acyl binding studies with the substrates [1-14C]acetyl-CoA and [2-14C]malonyl-CoA. Malonate was found to be transacylated to enzyme-bound pantetheine (SHc) exclusively by the Ser5421 hydroxyl group of malonyltransferase (OHMa1) while acetate could use both the acetyl (Ser819) and the malonyl (Ser5421) transferase active sites on its way to the SHc and SHp binding sites. Acylation of SHc with either substrate was unaffected by the absence of the 'peripheral' SH group (SHp) while binding of acetate to SHp was dependent on enzyme-bound pantetheine (SHc). These genetic data support a revised model regarding the intra-molecular channeling of acetate and malonate within yeast fatty acid synthase. Quantitative acyl binding studies revealed a maximum of 2-3 mol rather than the expected 12 mol of malonate and of 6-7 mol rather than 24 mol of acetate bound/mol hexameric yeast FAS. Only 20-30% of the malonyl-enzyme and 35-50% of the acetyl enzyme represented performic-acid-labile thioester bonds. The binding characteristics of both substrates, exhibiting Hill coefficients distinctly lower than 1, as well as their non-linear Lineweaver-Burk and Scatchard plots, point to a marked negative cooperativity among the 12 yeast FAS subunits. The observed sub-stoichiometric substrate binding characteristics of the enzyme are ascribed to this effect. An a priori asymmetry of the complex appears unlikely since the coenzyme-A:FAS transacylation equilibrium may be shifted towards the fully acetylated enzyme in the presence of N-ethylmaleimide. In contrast to the limited acylation capacity of the 'resting' enzyme, complete acylation of yeast FAS at all of its 12 SHc and SHp sites is observed under steady-state conditions of fatty acid biosynthesis. Under these conditions, the enzyme exhibits full-site reactivity at its SHp, SHc and OHAc sites, but a concomitant 18-fold increase in Km of the coenzyme-A:OHAc transacylation reaction keeps the acyl-O-ester content of the acylated enzyme at less than 5% of the total.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetates↗
PubMed1997

Comparison of the contractile effects and binding kinetics of endothelin-1 and sarafotoxin S6b in rat isolated renal artery.

1. To date, only two mammalian endothelin (ET) receptors, termed ETA and ETB, have been cloned, sequenced and characterized. However, several functional studies of isolated blood vessels suggest that ET-1-induced contractions may be mediated by multiple ETA receptors. In this study, the ETA receptors in renal arteries isolated from Wistar rats were characterized by isometric tension recording and radioligand binding techniques. 2. ET-1, sarafotoxin S6b (StxS6b) and ET-3 produced concentration-dependent contraction with similar response maxima in endothelium-denuded arteries, whereas the ETB receptor-selective agonist StxS6c was inactive. ET-1 and StxS6b were equipotent and 30 times more potent than ET-3. This agonist profile, together with the findings that the ETA receptor-selective antagonists, BQ-123 and FR-139317 caused concentration-dependent, rightward shifts of the concentration-effect curves to each agonist indicated that ET-1-induced contractions in rat renal artery were mediated via ETA receptors. 3. BQ-123 and FR-139317 were both significantly more potent inhibitors of contractions induced by StxS6b or ET-3 than of responses to ET-1, raising the possibility that a component of ET-1-induced contraction was mediated through atypical, BQ-123 (or FR-139317)-insensitive ETA receptors. However, in competition binding studies, specific [125I]-ET-1 and [125I]-StxS6b binding to rat renal artery sections was completely abolished by BQ-123 in a manner consistent with an action at a single site. Thus, competition binding studies did not provide any supportive evidence of the existence of a BQ-123-insensitive ETA receptor. 4. Additional studies revealed marked differences in the kinetics of [125I]-ET-1 and [125I]-StxS6b binding. Following a 3 h period of association of [125I]-ET-1 with its receptors, no significant dissociation of receptor-bound [125I]-ET-1 was observed during a 4 h washout period. In stark contrast, dissociation studies revealed that specific [125I]-StxS6b binding to ETA receptors was reversible (t0.5diss, 100 min). A series of association binding studies were also consistent with the specific binding of [125I]-ET-1 and [125I]-StxS6b being irreversible and reversible processes, respectively. 5. Thus, differences in BQ-123 potency against ET-1 and StxS6b-induced contractions in rat renal arteries might be due to differences in the kinetics of agonist binding, rather than due to the existence of atypical ETA receptors.

Animals↗
PubMed2005

Modeling binding kinetics at the Q(A) site in bacterial reaction centers.

Bacterial reaction centers (RCs) catalyze a series of electron-transfer reactions reducing a neutral quinone to a bound, anionic semiquinone. The dissociation constants and association rates of 13 tailless neutral and anionic benzo- and naphthoquinones for the Q(A) site were measured and compared. The K(d) values for these quinones range from 0.08 to 90 microM. For the eight neutral quinones, including duroquinone (DQ) and 2,3-dimethoxy-5-methyl-1,4-benzoquinone (UQ(0)), the quinone concentration and solvent viscosity dependence of the association rate indicate a second-order rate-determining step. The association rate constants (k(on)) range from 10(5) to 10(7) M(-)(1) s(-)(1). Association and dissociation rate constants were determined at pH values above the hydroxyl pK(a) for five hydroxyl naphthoquinones. These negatively charged compounds are competitive inhibitors for the Q(A) site. While the neutral quinones reach equilibrium in milliseconds, anionic hydroxyl quinones with similar K(d) values take minutes to bind or dissociate. These slow rates are independent of ionic strength, solvent viscosity, and quinone concentration, indicating a first-order rate-limiting step. The anionic semiquinone, formed by forward electron transfer at the Q(A) site, also dissociates slowly. It is not possible to measure the association rate of the unstable semiquinone. However, as the protein creates kinetic barriers for binding and releasing anionic hydroxyl quinones without greatly increasing the affinity relative to neutral quinones, it is suggested that the Q(A) site may do the same for anionic semiquinone. Thus, the slow semiquinone dissociation may not indicate significant thermodynamic stabilization of the reduced species in the Q(A) site.

Bacterial Proteins↗
PubMed1996

Immobilization of a recombinant cutinase by entrapment and by covalent binding. Kinetic and stability studies.

Fusarium solani pisi recombinant cutinase, immobilized by entrapment in calcium alginate and by covalent binding on porous silica, was used to catalyze the hydrolysis of tricaprylin. The influence of relevant parameters on the catalytic activity such as pH, temperature, and the substrate concentration were studied. Cutinase immobilized by entrapment presented a Michaelis-Menten kinetics for tricaprylin concentrations up to 200 mM. At higher concentrations of substrate, inhibition was observed. For covalent binding immobilization, diffusional limitations were observed at low substrate concentrations and substrate inhibition occurred for concentrations higher than 150 mM. The stability of immobilized cutinase was also evaluated. The enzyme immobilized by entrapment showed a high stability, in contrast to the immobilization on porous silica.

Caprylates↗
PubMed1994

The pteridine binding site of brain nitric oxide synthase. Tetrahydrobiopterin binding kinetics, specificity, and allosteric interaction with the substrate domain.

Nitric oxide (NO) synthases contain FAD, FMN, heme, and (6R)-5,6,7,8-tetrahydro-L-biopterin as prosthetic groups. We have characterized the pteridine-binding site of purified brain NO synthase, using 3H-labeled (6R)-5,6,7,8-tetrahydro-L-biopterin as radioligand. Association of [3H]tetrahydrobiopterin followed second-order kinetics (kon = 1.3 x 10(6) M-1 min-1), the dissociation reaction was reversible and first-order (koff = 3.2 x 10(-1) min-1), yielding a kinetic KD of 0.25 microM. Binding of the radioligand was competitively antagonized by several pteridine derivatives with the following order of potency (KI): 7,8-dihydro-L-biopterin (2.2 microM), (6S)-5,6,7,8-tetrahydro-L-biopterin (19 microM), (6R,S)-6-methyl-5,6,7,8-tetrahydropterin (240 microM), and 6,7-dimethyl-5,6,7,8-tetrahydropterin (> 1 mM). The affinity of NO synthase for tetrahydrobiopterin was increased 6-fold in the presence of 0.1 mM L-arginine (KD = 37 nM), and, conversely, tetrahydrobiopterin enhanced the affinity of the enzyme for 3H-labeled NG-nitro-L-arginine about 2-fold. 7-Nitroindazole, which presumably binds to the heme group of NO synthase, competitively inhibited binding of [3H]tetrahydrobiopterin and [3H]NG-nitro-L-arginine with similar Ki values (0.1 microM). Functional as well as binding studies revealed that 7-nitroindazole was competitive with both L-arginine and tetrahydrobiopterin. Our data indicate that brain NO synthase exhibits a highly specific binding site for (6R)-5,6,7,8-tetrahydro-L-biopterin, which allosterically interacts with the substrate domain and may be located proximal to the prosthetic heme group of NO synthase.

Allosteric Regulation↗
PubMed2002

Comparative analysis of cobalamin binding kinetics and ligand protection for intrinsic factor, transcobalamin, and haptocorrin.

Changes in the absorbance spectrum of aquo-cobalamin (Cbl x OH(2)) revealed that its binding to transcobalamin (TC) is followed by slow conformational reorganization of the protein-ligand complex (Fedosov, S. N., Fedosova, N. U., Nexø, E., and Petersen, T. E. (2000) J. Biol. Chem. 275, 11791-11798). Two phases were also observed for TC when interacting with a Cbl-analogue cobinamide (Cbi), but not with other cobalamins. The slow phase had no relation to the ligand recognition, since both Cbl and Cbi bound rapidly and in one step to intrinsic factor (IF) and haptocorrin (HC), namely the proteins with different Cbl specificity. Spectral transformations observed for TC in the slow phase were similar to those upon histidine complexation with Cbl x OH(2) and Cbi. In contrast to a closed structure of TC x Cbl x OH(2), the analogous IF and HC complexes revealed accessibility of Cbl's upper face to the external reagents. The binders decreased sensitivity of adenosyl-Cbl (Cbl x Ado) to light in the range: free ligand, IF x, HC x, TC x Cbl x Ado. The spectrum of TC x Cbl small middle dotAdo differed from those of IF and HC and mimicked Cbl x Ado participating in catalysis. The above data suggest presence of a histidine-containing cap shielding the Cbl-binding site in TC. The cap coordinates to certain corrinoids and, possibly, produces an incapsulated Ado-radical when Cbl small middle dotAdo is bound.

Binding Sites↗
PubMed1984

Binding kinetics of PAF-acether (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) to intact human platelets.

The binding of [3H]PAF-acether (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) to intact human gel-filtered platelets was measured at 22 degrees C. Specific binding reached saturation within 15 min at high doses of [3H]PAF-acether (0.5-0.9 nM), whereas about 90 min were required when low doses (0.02-0.5 nM) were used. Above 1 nM, [3H]PAF-acether non-specific binding increased progressively, which together with the demonstration of a 3H-labelled metabolite suggested uptake and metabolism of [3H]PAF-acether. Equilibrium analysis revealed one class of specific receptors with a Ka of 18.86 +/- 4.82 X 10(9) M-1 and 242 +/- 64 binding sites per platelet. Non-equilibrium binding revealed a similar Ka (16.87 X 10(9) M-1). Specific binding became irreversible after prolonged incubation, a process that was enhanced at increasing concentrations of [3H]PAF-acether. Platelets made desensitized to PAF-acether by prior incubation with unlabelled PAF-acether failed to bind a second dose of PAF-acether (3H-labelled), suggesting that desensitization resulted from loss of available binding sites. Under the conditions of the binding studies, PAF-acether induced exposure of the fibrinogen receptor, aggregation (in a stirred suspension) and alterations in (poly)-phosphatidylinositides. These results suggest that PAF-acether initiates platelet responses via receptor-mediated processes.

Adrenergic alpha-Antagonists↗
PubMed1987

Relationships between DNA-binding kinetics and biological activity for the 9-aminoacridine-4-carboxamide class of antitumor agents.

The kinetics of dissociation of calf thymus DNA complexes of the new intercalating antitumor drug N-[2-(dimethylamino)ethyl]-9-aminoacridine-4-carboxamide (5) and selected derivatives have been investigated by using the surfactant-sequestration method. The derivatives studied include those where the position (14 and 15) and nature of attachment (20 and 21) of the cationic side chain is modified, those where the distance (16-19) and composition (22-24) of the cationic group are varied, and those in which the chromophore is further substituted (25-31). While all of the compounds dissociate by a mechanism that involves at least three intermediate bound forms, derivatives bearing a 4-CONH(CH2)2NR1R2 side chain (where R1 and R2 are groups that permit the nitrogen to be protonated at neutral pH) have access to an additional binding mode of greater kinetic stability. A positive correlation is found between in vivo antitumor activity, selectivity of binding to GC-rich DNAs, and the presence of this fourth, long-lived transient species. We have interpreted our kinetic findings in terms of a molecular model for acridinecarboxamide-DNA complexes that accounts for the appearance of the fourth component. The acridine chromophore is postulated to intercalate from the narrow groove, its major axis lying at an angle to the major axis of the base pairs so that the CH atoms of positions 5 and 6 protrude into the groove. An important feature of the model is a bifurcated hydrogen bond between the O2 oxygen atom of a cytosine base adjacent to the binding site and the NH atoms of the carboxamide and protonated terminal amino functions of the drug molecule. Since the structural features required to form this bonding interaction are necessary, although not sufficient, conditions for in vivo antitumor activity, it is suggested that the model may describe the essential characteristics of the biologically active form of the bound drug. These findings further attest to the value of investigating the kinetics of DNA-drug interaction in studies of the mode of action of antitumor intercalating agents.

Amides↗
PubMed2003

A fractal analysis of protein to DNA binding kinetics using biosensors.

A fractal analysis of a confirmative nature only is presented for the binding of estrogen receptor (ER) in solution to its corresponding DNA (estrogen response element, ERE) immobilized on a sensor chip surface [J. Biol. Chem. 272 (1997) 11384], and for the cooperative binding of human 1,25-dihydroxyvitamin D(3) receptor (VDR) to DNA with the 9-cis-retinoic acid receptor (RXR) [Biochemistry 35 (1996) 3309]. Ligands were also used to modulate the first reaction. Data taken from the literature may be modeled by using a single- or a dual-fractal analysis. Relationships are presented for the binding rate coefficient as a function of either the analyte concentration in solution or the fractal dimension that exists on the biosensor surface. The binding rate expressions developed exhibit a wide range of dependence on the degree of heterogeneity that exists on the surface, ranging from sensitive (order of dependence equal to 1.202) to very sensitive (order of dependence equal to 12.239). In general, the binding rate coefficient increases as the degree of heterogeneity or the fractal dimension of the surface increases. The predictive relationships presented provide further physical insights into the reactions occurring on the biosensor surface. Even though these reactions are occurring on the biosensor surface, the relationships presented should assist in understanding and in possibly manipulating the reactions occurring on cellular surfaces.

Biosensing Techniques↗
PubMed2002

Determination of drug-plasma protein binding kinetics and equilibria by chromatographic profiling: exemplification of the method using L-tryptophan and albumin.

Drug-plasma protein binding may greatly influence the bioavailability and metabolism of a plasma-borne drug, the bound form being partially protected from the metabolic fate of the unbound drug. Traditionally, equilibrium values (e.g., percentage binding) for drug-protein binding have been measured to rationalize in vivo phenomena. However, such studies overlook the influence of kinetics. A rapid method of simultaneously determining kinetic rate constants and equilibrium constants from chromatographic profiles has been developed, based on the use of immobilized protein columns and HPLC. By measuring the chromatographic profiles (the position and width) of a retained and an unretained compound one can directly determine both the rate and equilibrium constants. Results are presented for the binding of L-tryptophan to human serum albumin to exemplify the method. The association equilibrium constant (Ka) and the association and dissociation rate constants (k(a) and k(d), respectively) were thereby measured in an aqueous pH 7.4 environment at 37 degrees C as 0.84 10(4) M(-1), 5.8 10(4) M(-1) s(-1), and 6.9 s(-1), respectively. These compare favorably with previously published results. The described method may be used in quantitative structure-property relationship-based rational drug discovery or for the rationalization of drug pharmacokinetics.

Animals↗
PubMed2000

Dioxygen-binding kinetics and thermodynamics of a series of dicopper(I) complexes with bis[2-(2-pyridyl)ethyl]amine tridendate chelators forming side-on peroxo-bridged dicopper(II) adducts.

Copper-dioxygen interactions are of interest due to their importance in biological systems as reversible O2- carriers, oxygenases, or oxidases and also because of their role in industrial and laboratory oxidation processes. Here we report on the kinetics (stopped-flow, -90 to 10 degrees C) of O2-binding to a series of dicopper(I) complexes, [Cu2(Nn)(MeCN)2]2+ (1Nn) (-(CH2)n- (n = 3-5) linked bis[(2-(2-pyridyl)ethyl]amine, PY2) and their close mononuclear analogue, [(MePY2)Cu(MeCN)]+ (3), which form mu-eta 2:eta 2-peroxodicopper(II) complexes [Cu2(Nn)-(O2)]2+ (2Nn) and [(MePY2)Cu]2(O2)]2+ (4), respectively. The overall kinetic mechanism involves initial reversible (k+,open/k-,open) formation of a nondetectable intermediate O2-adduct [Cu2(Nn)(O2)]2+ (open), suggested to be a CuI...CuII-O2- species, followed by its reversible closure (k+,closed/k-,closed) to form 2Nn. At higher temperatures (253 to 283 K), the first equilibrium lies far to the left and the observed rate law involves a simple reversible binding equilibrium process (kon,high = (k+,open/k-,open)(k+,closed)). From 213 to 233 K, the slow step in the oxygenation is the first reaction (kon,low = k+,open), and first-order behavior (in 1Nn and O2) is observed. For either temperature regime, the delta H++ for formation of 2Nn are low (delta H++ = -11 to 10 kJ/mol; kon,low = 1.1 x 10(3) to 4.1 x 10(3) M-1 s-1, kon,high = 2.2 x 10(3) to 2.8 x 10(4) M-1 s-1), reflecting the likely occurrence of preequilibria. The delta H degree ranges between -81 and -84 kJ mol-1 for the formation of 2Nn, and the corresponding equilibrium constant (K1) increases (3 x 10(8) to 5 x 10(10) M-1; 183 K) going from n = 3 to 5. Below 213 K, the half-life for formation of 2Nn increases with, rather than being independent of, the concentration of 1Nn, probably due to the oligomerization of 1Nn at these temperatures. The O2 reaction chemistry of 3 in CH2Cl2 is complicated, including the presence of induction periods, and could not be fully analyzed. However, qualitative comparisons show the expected slower intermolecular reaction of 3 with O2 compared to the intramolecular first-order reactions of 1Nn. Due to the likelihood of the partial dimerization of 3 in solution, the t1/2 for the formation of 4 remains constant with increasing complex concentration rather than decreasing. Acetonitrile significantly influences the kinetics of the O2 reactions with 1Nn and 3. For 1N4, the presence of MeCN inhibits the formation of a previously (Jung et al, J. Am. Chem. Soc. 1996, 118, 3763-3764) observed intermediate. Small amounts of added MeCN considerably slow the oxygenation rates of 3, inhibit its full formation to 4, and increase the length of the induction period. The results for 1Nn and their mononuclear analogue 3 are presented, and they are compared with each other as well as with other dinucleating dicopper(I) systems.

Copper↗
PubMed1981

Binding kinetics of methyl alpha-D-mannopyranoside to concanavalin A: temperature-jump relaxation study with 4-methylumbelliferyl alpha-D-mannopyranoside as a fluorescence indicator ligand.

The binding of methyl alpha-D-mannopyranoside and methyl alpha-D-glucopyranoside to concanavalin A has been investigated by the temperature-jump relaxation kinetic technique using the competitive inhibitor 4-methylumbelliferyl alpha-D-mannopyranoside as an indicator of the binding reaction. The analysis shows that these saccharides bind to concanavalin A in a single bimolecular step. The binding parameters are compared to those of derivatized carbohydrates which have previously been used to study the binding of saccharides to concanavalin A. The similarity of the association rate constants indicates that a common process is involved in the binding of all carbohydrates to concanavalin A. The different affinities of saccharides for the lectin are primarily due to the different dissociation rate constants. A discussion of the proposed mechanism is given under the Appendix to clarify the fact that one of the observed relaxation times is faster than is possible with only the kinetic indicator reaction.

Binding, Competitive↗
PubMed2000

Equilibrium and kinetic binding interactions between DNA and a group of novel, nonspecific DNA-binding proteins from spores of Bacillus and Clostridium species.

Binding of alpha/beta-type small acid-soluble spore proteins (SASP) is the major determinant of DNA resistance to damage caused by UV radiation, heat, and oxidizing agents in spores of Bacillus and Clostridium species. Analysis of several alpha/beta-type SASP showed that these proteins have essentially no secondary structure in the absence of DNA, but become significantly alpha-helical upon binding to double-stranded DNAs or oligonucleotides. Folding of alpha/beta-type SASP induced by a variety of DNAs and oligonucleotides was measured by CD spectroscopy, and this allowed determination of a DNA binding site size of 4 base pairs as well as equilibrium binding parameters of the alpha/beta-type SASP-DNA interaction. Analysis of the equilibrium binding data further allowed determination of both intrinsic binding constants (K) and cooperativity factors (omega), as the alpha/beta-type SASP-DNA interaction was significantly cooperative, with the degree of cooperativity depending on both the bound DNA and the salt concentration. Kinetic analysis of the interaction of one alpha/beta-type SASP, SspC(Tyr), with DNA indicated that each binding event involves the dimerization of SspC(Tyr) monomers at a DNA binding site. The implications of these findings for the structure of the alpha/beta-type SASP.DNA complex and the physiology of alpha/beta-type SASP degradation during spore germination are discussed.

Bacillus↗
PubMedDate not supplied

Tissue binding kinetics of tobramycin. An experimental study in the mouse inner ear in vitro.

The tissue-binding capacity of tritiated tobramycin (TM) was analysed in an organ culture system in the embryonic inner ear of the mouse. A rapid and probably irreversible binding of TM occurs within 10 min, showing about 100 micrograms TM per gram protein in the vestibular half and about 75 micrograms TM per gram protein in the cochlear half of the labyrinth. A steady state is reached within 60 min. The TM uptake was then 390 micrograms TM per gram protein in the vestibular part and 270 micrograms in the cochlear part of the inner ear. A minor fraction of TM, of the same magnitude as initially bound, becomes irreversibly bound, whereas the slow accumulation which reaches the steady state level seems reversible.

Animals↗