Slow Conformational Change Research Articles

Activation enthalpies and pH dependence of phenol hydroxylase from Trichosporon cutaneum, in vitro and in situ

The effect of pH and temperature on phenol hydroxylase in vitro was compared to the corresponding effect on the enzyme in situ, in permeabilized cells. Activation enthalpies in situ were about 75–80% of those in vitro, in both cases decreasing with increasing pH (6.0–8.5). The order of addition of phenol and NADPH affected the K m values for phenol at 25°C, but not at 10°C. The results support the idea that the enzyme in situ is in a more favourable position for catalysis than the purified enzyme and that slow conformational changes, triggered by binding of phenol, become rate limiting above 10°C.

Binding of Ca2+ to the calcium adenosinetriphosphatase of sarcoplasmic reticulum.

The binding of Ca2+ and the resulting change in catalytic specificity that allows phosphorylation of the calcium ATPase of sarcoplasmic reticulum by ATP were examined by measuring the amount of phosphoenzyme formation from [32P]ATP, or 45Ca incorporation into vesicles, after the simultaneous addition of ATP and EGTA at different times after mixing enzyme and Ca2+ (25 degrees C, pH 7.0, 5 mM MgSO4, 0.1 M KCl). A "burst" of calcium binding in the presence of high [Ca2+] gives approximately 12% phosphorylation and internalization of two Ca2+ at very short times after the addition of Ca2+ with this assay. This shows that calcium binding sites are available on the cytoplasmic-facing side of the free enzyme. Calcium binding to these sites induces the formation of cE.Ca2, the stable high-affinity form of the enzyme, with k = 40 s-1 at saturating [Ca2+] and a half-maximal rate at approximately 20 microM Ca2+ (from Kdiss = 7.4 X 10(-7) M for Ca.EGTA). The formation of cE.Ca2 through a "high-affinity" pathway can be described by the scheme E 1 in equilibrium cE.Ca1 2 in equilibrium cE.Ca2, with k1 = 3 X 10(6) M-1 s-1, k2 = 4.3 X 10(7) M-1 s-1, k-1 = 30 s-1, k-2 = 60 s-1, K1 = 9 X 10(-6) M, and K2 = 1.4 X 10(-6) M. The approach to equilibrium from E and 3.2 microM Ca2+ follows kobsd = kf + kr = 18 s-1 and gives kf = kr = 9 s-1. The rate of exchange of 45Ca into the inner position of cE.Ca2 shows an induction period and is not faster than the approach to equilibrium starting with E and 45Ca. The dissociation of 45Ca from the inner position of cE.45Ca.Ca in the presence of 3.2 microM Ca2+ occurs with a rate constant of 7 s-1. These results are inconsistent with a slow conformational change of free E to give cE, followed by rapid binding-dissociation of Ca2+.

Enzymes as biosensors. 1. Enzyme memory and sensing chemical signals.

When a free enzyme exists under different conformations that 'slowly' isomerize during the conversion of a substrate into a product, the corresponding 'slow' relaxation component may interfere with the steady-state component. The apparent steady-state rate that may be measured under these conditions is called the meta-steady-state rate for it refers to the existence of metastable states of the enzyme during the reaction. By contrast to the real steady-state rate, the meta-steady-state rate is dependent upon the initial state of the enzyme, that is on the respective concentrations of the free enzyme forms. The simplest model that may display this type of behaviour is the mnemonical model. For a fixed concentration of the last product of the reaction sequence the meta-steady state is different depending on that concentration being reached by an increase or a decrease of a previous concentration. This means that the meta-steady-state rate describes a hysteresis loop as the product concentration is increased and decreased. Owing to the existence of metastable states, the enzyme system behaves as a biosensor that is able to detect both a concentration and the direction of a concentration change. The existence of the hysteresis loop of the meta-steady-state rate implies that the two free enzyme forms display hysteresis as well. A chemical potential, called the sensing potential, is specifically associated with the 'perception' of the direction of the thermodynamic force generated by the decrease or the increase of the concentration of the ligand that binds to one of the enzyme conformations. The sensing potential of the enzyme conformer that does not bind the product increases and reaches a plateau as the chemical potential of that product is raised. Alternatively the sensing potential of the other conformer vanishes at low and high chemical potentials of the product and is significant for intermediate chemical potentials. Enzymes that display very slow conformation changes may thus be viewed as elementary sensor devices.

A slow kinetic transient in RNA synthesis catalysed by wheat-germ RNA polymerase II

Progress curves of U-A-primed RNA synthesis catalysed by wheat-germ RNA polymerase II on a poly[d(A-T)] template exhibit a slow burst of activity. In contrast, the progress curves of single-step addition of UMP to U-A primer in the abortive elongation reaction do not exhibit the slow burst of activity. The correlation between the kinetic transient in the productive pathway of RNA synthesis and the rate of abortive elongation is suggestive of the occurrence of a slow conformational change of the transcription complex during the transition from abortive to productive elongation. The exceptional duration of the transient burst (in the region of 4 min) may suggest a transition of a hysteretic type.

Thiol- and pH-modulated slow conformational changes and cooperativity of phenol-binding sites in phenol hydroxylase.

Spectrophotometric titration of phenol hydroxylase (EC 1.14.13.7) with phenol indicated interacting sites for phenol binding. In the absence of added thiol, the cooperativity was positive up to a pH around 8.0 but negative at higher pH values. With added thiol-ethylenediaminetetraacetate, the cooperativity was negative at all investigated pH values. Conversely, a corresponding titration of an enzyme preparation that had been selectively modified in its two most reactive SH groups indicated positive cooperativity at all studied pH values. This selective modification affects the activity of the enzyme to a very minor degree, in contrast to more extensive SH blocking, which displaces flavin adenine dinucleotide with a corresponding loss of activity [Neujahr, H. Y., & Gaal, A. (1975) Eur. J. Biochem. 58, 351-357]. The reactivity of SH groups in the enzyme was significantly decreased after turnover. Thiol treatment restored it to that of the native enzyme. Adding phenol prior to reduced nicotinamide adenine dinucleotide phosphate (NADPH) in the assay of phenol hydroxylase gave immediate linearity and higher initial rates than when NADPH was added first. In the absence of added thiol, there was then a shift of the pH optimum. The results indicate slow conformational changes limiting the rate of the overall reaction. The two most reactive SH groups of phenol hydroxylase, though not participating in any obvious redox reactions, are important for these slow conformational changes and for the cooperativity of phenol-binding sites, wherein the anionic S- forms may be involved (pKa for cysteine is 8.35).

Reactions of the sarcoplasmic reticulum calcium adenosinetriphosphatase with adenosine 5'-triphosphate and Ca2+ that are not satisfactorily described by an E1-E2 model.

Phosphorylation of the sarcoplasmic reticulum calcium ATPase, E, is first order with kb = 70 +/- 7 s-1 after free enzyme was mixed with saturating ATP and 50 microM Ca2+; this is one-third the rate constant of 220 s-1 for phosphorylation of enzyme preincubated with calcium, cE.Ca2, after being mixed with ATP under the same conditions (pH 7.0, Ca2+-loaded vesicles, 100 mM KCl, 5 mM Mg2+, 25 degrees C). Phosphorylation of E with ATP and Ca2+ in the presence of 0.25 mM ADP gives approximately 50% E approximately P.Ca2 with kobsd = 77 s-1, not the sum of the forward and reverse rate constants, kobsd = kf + kr = 140 s-1, that is expected for approach to equilibrium if phosphorylation were rate limiting. These results show that (1) kb represents a slow conformational change, rather than phosphoryl transfer, and (2) different pathways are followed for the phosphorylation of E and of cE.Ca2. The absence of a lag for phosphorylation of E with saturating ATP and Ca2+ indicates that all other steps, including the binding of Ca2+ ions and phosphoryl transfer, have rate constants of greater than 500 s-1. Chase experiments with unlabeled ATP or with ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) show that the rate constants for dissociation of [gamma-32P]ATP and Ca2+ are comparable to kb. Dissociation of ATP occurs at 47 s-1 from E.ATP.Ca2+ and at 24 s-1 from E.ATP. Approximately 20% phosphorylation occurs following an EGTA chase 4.5 ms after the addition of 300 microM ATP and 50 microM Ca2+ to enzyme. This shows that Ca2+ binds rapidly to the free enzyme, from outside the vesicle, before the conformational change (kb). The fraction of Ca2+-free E.[gamma-32P]ATP that is trapped to give labeled phosphoenzyme after the addition of Ca2+ and a chase of unlabeled ATP is half-maximal at 6.8 microM Ca2+, with a Hill slope of n = 1.8. The calculated dissociation constant for Ca2+ from E.ATP.Ca2 is approximately 2.2 X 10(-10) M2 (K0.5 = 15 microM). The rate constant for the slow phase of the biphasic reaction of E approximately P.Ca2 with 1.1 mM ADP increases 2.5-fold when [Ca2+] is decreased from 50 microM to 10 nM, with half-maximal increase at 1.7 microM Ca2+. This shows that Ca2+ is dissociating from a different species, aE.ATP.Ca2, that is active for catalysis of phosphoryl transfer, has a high affinity for Ca2+, and dissociates Ca2+ with k less than or equal to 45 s-1.(ABSTRACT TRUNCATED AT 400 WORDS)

A stopped-flow study of the reaction of cytochrome c peroxidase with hydroperoxides

Transient kinetic measurements show that cytochrome c peroxidase reacts with excess of hydroperoxides to produce compound ES in two phases. The activation energies for the fast and slow phases are calculated to be 6.3 and 20.5 kcal·mol −1, respectively. The fast phase is assigned to the reaction of native active (pulsed) cytochrome c peroxidase with peroxides, whereas the slow phase is due to the presence of an inactive (aged, resting) enzyme. As the active species is exhausted, the equilibrium between the active and inactive enzymes is shifted by a slow conformational change to replenish the active enzyme. Since the rate-limiting step of the reaction of the inactive enzyme with peroxides is the conformation change, the overall reaction rate is independent of the nature and concentration of peroxides.

Formula omitted] kinetics of phenol hydroxylase

The half saturation constant for phenol was much lower with phenol hydroxylase in situ than with the purified enzyme, whereas the constant for NADPH was higher. In both cases, the linearized plots of the Michaelis-Menten equation were biphasic and the half saturation constants for all phenolic substrates were several times lower, when the phenol was added to the assay medium before NADPH, than when NADPH was added first. There was a similar, but much smaller, effect on the half saturation constants for NADPH. The V-values were not affected by the order of addition. The results suggest slow conformational changes in the enzyme during the overall reaction, which seem even slower, when the enzyme is measured in situ .

Evidence that the slow conformation change controlling NADH release from the enzyme is rate-limiting during the oxidation of propionaldehyde by aldehyde dehydrogenase.

The displacement of NADH from the aldehyde dehydrogenase X NADH complex by NAD+ was followed at pH 7.0, and the data were fitted by a non-linear least-squares iterative procedure. At pH 7.0 the decay constants for the dissociation of NADH from aldehyde dehydrogenase X NADH complexes (1.62 +/- 0.09 s-1 and 0.25 +/- 0.004 s-1) were similar to the values previously determined by MacGibbon, Buckley & Blackwell [(1977) Biochem. J. 165, 455-462] at pH 7.6, and apparent differences between these values and those reported by Dickinson [(1985) Biochem. J. 225, 159-165] are resolved. Experiments at low concentrations of propionaldehyde show that isomerization of a binary E X NADH complex is part of the normal catalytic mechanism of the enzyme. Evidence is presented that the active-site concentration of aldehyde dehydrogenase is halved when enzyme is pre-diluted to low concentrations before addition of NAD+ and substrate. The consequences of this for the reported values of kcat. are discussed. A general mechanism for the aldehyde dehydrogenase-catalysed oxidation of propionaldehyde which accounts for the published kinetic data, at concentrations of aldehyde which bind only at the active site, is presented.

Characteristics of the formation of enzyme-bound ATP from medium inorganic phosphate by mitochondrial F1 adenosinetriphosphatase in the presence of dimethyl sulfoxide.

Addition of dimethyl sulfoxide promotes the formation of enzyme-bound ATP from medium Pi by mitochondrial F1 adenosinetriphosphatase that has tightly bound ADP present. Measurements are reported of medium Pi in equilibrium H18OH exchange and of the dependence of formation of enzyme-bound ATP on Pi concentration. Attainment of an apparent equilibrium between medium Pi and bound ATP requires longer than 30 min, even though the rates of Pi binding and release after apparent equilibrium is reached would suffice for a faster approach to equilibrium. Slow protein conformational changes or other unknown modulating factors may be responsible for the slow rate of bound ATP formation. After apparent equilibrium is reached, each Pi that binds to the enzyme reversibly forms ATP about 50 times before being released to the medium. The rate of interconversion of bound ATP to bound ADP and Pi is much slower than that in the absence of dimethyl sulfoxide as measured with sufficiently low ATP concentrations so that single-site catalysis is favored. Although the interconversion rate is slowed, the equilibrium constant for bound ATP formation from bound ADP and Pi is not far from unity. Dimethyl sulfoxide favors the formation of enzyme-bound ATP by promoting the competent binding of Pi to enzyme with ADP bound at a catalytic site rather than by promoting formation of bound ATP from bound ADP and Pi.

The effects of Mg2+ at the high-affinity and low-affinity sites on the polymerization of actin and associated ATP hydrolysis.

Actin contains a single high-affinity cation-binding site, for which Ca2+ and Mg2+ can compete, and multiple low-affinity cation-binding sites, which can bind Ca2+, Mg2+, or K+. Binding of cations to the low-affinity sites causes polymerization of monomeric actin with either Ca2+ or Mg2+ at the high-affinity site. A rapid conformational change occurs upon binding of cations to the low-affinity sites (G----G) which is apparently associated with the initiation of polymerization. A much slower conformational change (G----G', or G----G' if the low-affinity sites are also occupied) follows the replacement of Ca2+ by Mg2+ at the high-affinity site. This slow conformational change is reflected in a 13% increase in the fluorescence of G-actin labeled with the fluorophore 7-chloro-4-nitrobenzene-2-oxadiazole (NBD-labeled actin). The rate of the ATP hydrolysis that accompanies elongation is slower with Ca-G-actin than with Mg-G'-actin (i.e. with Ca2+ rather than Mg2+ at the high-affinity site) although their rates of elongation are similar. The slow ATP hydrolysis on Ca-F-actin causes a lag in the increase in fluorescence associated with the elongation of actin labeled with the fluorophore N-pyrene iodoacetamide (pyrenyl-labeled actin), even though there is no lag in the elongation rate, because pyrenyl-labeled ATP-F-actin subunits have a lower fluorescence intensity than pyrenyl-labeled ADP-F-actin subunits. The effects of the cation bound to the high-affinity binding site must, therefore, be considered in quantitatively analyzing the kinetics of polymerization of NBD-labeled actin and pyrenyl-labeled actin. Although their elongation rates are not very different, the rate of nucleation is much slower for Ca-G-actin than for Mg-G'-actin, probably because of the slower rate of ATP hydrolysis when Ca2+ is bound to the high-affinity site.

Relationship between the affinity and proteolysis of the insulin receptor. Evidence that higher affinity receptors are preferentially degraded.

125I-Insulin binding to rat liver plasma membranes initiated two processes that occurred with similar time courses: an increase of receptor affinity for hormone and degradation of the Mr 135,000 alpha subunit of the insulin receptor to a fragment of Mr 120,000. Inhibitors of serine proteinases prevented alpha subunit degradation without affecting the affinity change. This shows that the change of affinity is not produced by receptor proteolysis and that the intact alpha subunit of the insulin receptor can exist as a higher or lower affinity species. Hormone binding was much more rapid than receptor proteolysis and the initial rate of alpha subunit degradation was independent of the concentration of occupied lower affinity receptors. Only persistent hormone binding and the accumulation of higher affinity insulin-receptor complexes led to significant receptor proteolysis. As the incubation time between 125I-insulin and membranes increased, the rate at which hormone dissociated from Mr 135,000 complexes diminished, whereas hormone dissociated from Mr 120,000 complexes slowly after brief or extended incubations. These observations suggest that 125I-insulin binds to membranes to form low affinity complexes that are not substrates for proteolysis. A slow conformational change produces higher affinity hormone-receptor complexes that are selectively degraded. Thus, the conversion between states of affinity may play a role in the regulation of receptor proteolysis and, consequently, insulin action in cells.

Protein binding to two-dimensional hydrophobic binding-site lattices: Sorption kinetics of phosphorylase b on immobilized butyl residues

The adsorption and desorption kinetics of the binding of the large ( m = 200 kDa) protein, phosphorylase b, to a hydrophobic surface (butyl agarose) was studied. The initial adsorption rate is a hyperbofic function of the initial free protein concentration (saturation kinetics). At 5°C the apparent dissociation constant for the half-maximal rate ( K d ) is 13.2 μ M. A maximal rate ( V max) of 43 μmole/(liter packed agarose × min) is obtained. The first-order rate constant of the rate-limiting step of adsorption ( k 2) is 1.8 × 10 −3 s −1 ( t 1 2 ∼ 6 min) and illustrates that the adsorption of this protein to a hydrophobic surface is a slow process. It is concluded that adsorption is a multistep reaction and that the rate-limiting step is not the collision rate but a slow conformational change or reorientation step of the protein on the butyl agarose surface. From the initial desorption rates, measured in dilution experiments, the apparent, first-order off-rate constants k′ off) were determined. It is shown that the basal, off-rate constant ( k′ off ∼ 64 × 10 −6 s −1; t 1 2 ∼ 180 min) obtained with butyl agarose having a surface concentration ca. 320 nmole butyl residues/m 2, does not show the constancy expected for the binding of a protein to single, independent sites on a surface. In fact, the off-rate constant depends on the fractional saturation of the adsorbent surface with protein and is sensitive to the presence of other protein(s) in the bulk. It is concluded that here is one of the first examples of a protein showing negative cooperative surface kinetics on an adsorbent.

Conformations of fibroblast and E. coli-derived recombinant human interferon-beta s as studied by nuclear magnetic resonance and circular dichroism.

The conformations of fibroblast and E. coli-derived recombinant human interferon-beta s were studied by circular dichroism and nuclear magnetic resonance spectroscopy in the acidic pH region of 4.6 to 1.6. Both interferons have very similar conformations with high alpha-helix contents (approximately 70%). These results suggest that glycosylation does not appreciably change the conformation of human interferon-beta. Moreover, a slow conformational change is observed below pH 2.0, which induces the disruption of beta-sheets.

Kinetics of the interaction of N-(phosphonacetyl)-L-aspartate with the catalytic subunit of aspartate transcarbamoylase. A slow conformational change subsequent to binding.

The binding of the bisubstrate ligand N-(phosphonacetyl)-L-aspartate (PALA) to the active sites of both the free catalytic subunit of aspartate transcarbamoylase and the intact holoenzyme causes conformational changes which have been studied extensively. However, no kinetic information has been available about the sequence of events occurring during the formation or dissociation of the complexes. Stopped flow kinetics, 31P saturation transfer NMR spectroscopy, and presteady-state kinetics were used to monitor the interaction of PALA with the catalytic subunit (or a derivative containing nitrotyrosyl chromophores which served as spectral probes). The various experimental approaches lead to a mechanism that includes a rapid binding of PALA with an "on" rate of about 10(8)M-1s-1 and an "off" rate of 28 s-1, followed by a much slower isomerization of the complex with a forward rate constant of 0.18 s-1. Analysis of the presteady-state bursts of enzyme activity when the protein is added to a mixture of substrates and PALA and of the lag in activity when the PALA complex with catalytic subunit is added to substrates yielded a rate constant for the reverse isomerization of 0.018s-1. Thus, the conformational change subsequent to PALA binding leads to a 10-fold increase in the equilibrium constant for complex formation. Stopped flow kinetic measurements of the spectral change resulting from mixing the complex of PALA and nitrated protein with native enzyme showed a slow process with a t1/2 of about 11 s, whereas 31P saturation transfer NMR experiments yielded at t1/2 of about 260 ms for the dissociation of PALA from the complex. This apparent disparity is understood in terms of the two-step binding scheme where rapid dissociation of the initial ligand X enzyme complex is measured by the NMR technique and the slow isomerization of the complex is responsible for the bulk of the stopped flow signal.

Structure and Dynamics of Neurotoxins from Snake Venoms as Revealed by Nuclear Magnetic Resonance and Circular Dichroism

AbstractShort and long neurotoxins from snake venoms were studied by nuclear magnetic resonance (NMR) and circular dichroism (CD). From the analysis of amide and a proton resonances of Laticauda semifasciata III, triple stranded antiparallel pleated β-sheet structure was found to be stable even in aqueous solution. The NMR analysis showed that this α-sheet is common structural feature to erabutoxin b, α-nigricolis, α-cobratoxin and α-bungarotoxin. Since amino acid rersidues on this β-sheet are essential for neurotoxicity, this α-sheet is supposed to be the binding surface to the acetylcholine receptor proteins. In contrast to the crystal structure of proteins, proteins in aqueous solution are more mobile and respond to the change of the environment. pH and temperature dependence of NMR spectra of α-cobratoxin was studied by NMR and CD. Detailed analysis of NMR spectra showed that protonation of His-18 disrupts the hydrophobic core which induces slow conformation change, while the loop part is sensitive to...

Interaction between the mitochondrial ATP synthetase and ATPase inhibitor protein: Active/inactive slow pH-dependent transitions of the inhibitor protein

The rate of mitochondrial ATPase inactivation by the naturally occurring inhibitor protein in the presence of saturating ATP and Mg 2+ at pH 8.0 depends hyperbolically on the amount of inhibitor added; the upper limit of an apparent first-order constant for the inactivation process is 1.0 −1 at 25°C. A dramatic difference in the inactivation rate is observed when the protein inhibitor is added to the same assay system from either acidic (pH 4.8) or alkaline (pH 8.2) solutions. The slow reversible transition of the inhibitor from its rapidly reacting ‘acidic’ form to the slow reacting ‘alkaline’ form occurs when the solution of the protein inhibitor is subjected to a pH-jump from 4.8 to 8.2 ( t 1 2 ~ 30 s at 25°C). The pH-profile of the inhibitor active/inactive equilibrium suggests that a group with p K a ~ 6.5 is involved in the transition. Treatment of the inhibitor protein with a histidine-specific reagent (e.g. diethyl pyrocarbonate) abolishes its inactivating effect on the ATPase activity. It is concluded that the protonation/deprotonation of the inhibitor protein followed by its slow conformational changes is the rate-limiting step in the inhibitor-ATP synthetase interaction.

The 'slow' pH-induced conformational transition of chloroplast fructose 1,6-bisphosphatase and the control of the Calvin cycle.

A slow conformation change of chloroplastic reduced fructose bisphosphatase is detected upon raising the pH from 7 to 8 or upon lowering the pH from 8 to 7. These conformation changes are fully reversible. In the time scale investigated these processes occur in one step. Their time constants and their amplitudes have been determined and analyzed as a function of proton concentration. The results obtained are consistent with the view that upon ionization or protonation of a strategic ionizable group the protein undergoes a 'slow' conformational transition that may be followed by conventional fluorescence techniques. Since illumination brings about a pH rise of chloroplastic stroma from 7 to 8, the above results suggest that light activation of fructose bisphosphatase is at least in part due to a slow conformation change of this enzyme.

Self-association of bovine prothrombin fragment 1 in the presence of metal ions. Use of a covalent cross-linking reagent to study the reaction.

The interaction of metal ions with prothrombin fragment 1 has been shown by other investigators to result in conformational change(s). Previous studies based on hydrodynamic measurements from other laboratories have suggested that prothrombin fragment 1 will, in addition to undergoing conformational change, self-associate in the presence of metal ions but there is a question regarding the specificity and role of metal ions in this process. The present study has made use of a covalent cross-linking agent, dithiobis(succinimidylpropionate), to study the self-association of prothrombin fragment 1 and has focused on factors influencing the covalent dimerization of bovine prothrombin fragment 1 during reaction with this reagent. Optimal dimerization of bovine prothrombin fragment 1 required the involvement of cations in two separable processes. The "first role" is probably a reflection of a slow conformational change in the bovine prothrombin fragment 1 representing isomerization of a proline-containing peptide bond (Marsh, H. C., Scott, M. E., Hiskey, R. G., and Koehler, K. A. (1979) Biochem. J. 183, 513-517). The "second role" most likely reflects ion bridging with another prothrombin fragment 1 molecule. The ion specificities of these two roles are clearly different. Ions stabilizing the proper conformation (first role) are Ca2+, Sr2+, Mg2+, Mn2+, with Ba2+ having less capability in filling this role. Ions which have activity in the second role (ion bridge formation) are Ca2+, Sr2+, and Ba2+. Mn2+ and Mg2+ cannot fulfill the requirements of this second role and actually inhibit the function of Ca2+.

Erratum: Transient kinetics of chemical reactions with bounded diffusion perpendicular to the reaction coordinate: Intramolecular processes with slow conformational changes [J. Chem Phys. 7 8, 6947 (1983)

Erratum: Transient kinetics of chemical reactions with bounded diffusion perpendicular to the reaction coordinate: Intramolecular processes with slow conformational changes [J. Chem Phys. <b>7</b> <b>8</b>, 6947 (1983)