Relative association of Rubisco with manganese and magnesium as a regulatory mechanism in plants

Rubisco, the enzyme that constitutes as much as half of the protein in a leaf, initiates either the photorespiratory pathway that supplies reductant for the assimilation of nitrate into amino acids or the C3 carbon fixation pathway that generates carbohydrates. The relative rates of these two pathways depend both on the relative extent to which O₂ and CO₂ occupies the active site of Rubisco and on whether manganese or magnesium is bound to the enzyme. This study quantified the activities of manganese and magnesium in isolated tobacco chloroplasts and the thermodynamics of binding of these metals to Rubisco purified from tobacco or a bacterium. In tobacco chloroplasts, manganese was less active than magnesium, but Rubisco purified from tobacco had a higher affinity for manganese. The activity of each metal in the chloroplast was similar in magnitude to the affinity of tobacco Rubisco for each. This indicates that, in tobacco chloroplasts, Rubisco associates almost equally with both metals and rapidly exchanges one metal for the other. Binding of magnesium was similar in Rubisco from tobacco and a bacterium, whereas binding of manganese differed greatly between the Rubisco from these species. Moreover, the ratio of leaf manganese to magnesium in C3 plants increased as atmospheric CO₂ increased. These results suggest that Rubisco has evolved to improve the energy transfers between photorespiration and nitrate assimilation and that plants regulate manganese and magnesium activities in the chloroplast to mitigate detrimental changes in their nitrogen/carbon balance as atmospheric CO₂ varies.     

Theoretical study on phthalocyanine,pyrazinoporphyrazine and their complexation with Mg2+ and Zn2+

We have studied the UV–vis absorption spectra of metal-free phthalocyanine (H₂Pc), metal-free pyrazinoporphyrazine (H₂PyzPz) and their complexes with Mg^2 +  and Zn^2 +  using semiempirical Zerners intermediate neglect of differential overlap and time-dependent density functional theory methods. The predicted absorption spectra of H₂Pc and their complexes are in agreement with a previous experiment report. The calculated results show that the Q band absorption peaks of H₂PyzPz and their complexes are blue-shifted by 40 nm as compared with those of H₂Pc and their complexes, respectively. The frontier molecular orbitals (HOMO and LUMO) of H₂Pc, H₂PyzPz and their metal complexes were investigated as well. The nitrogen atoms in the pyrazine rings stabilise the HOMO more than the LUMO, and the deprotonation of the pyrrole rings induced by the metal coordination destabilises the LUMO more than the HOMO. Because of the increased band gap, the absorption bands of H₂PyzPz and the metal-coordinated compounds are blue-shifted.     

Molecular basis for the integration of inositol phosphate signaling pathways via human ITPK1

The synthesis and the metabolism of inositol 3,4,5,6-tetrakisphosphate (Ins(3,4,5,6)P₄) are the responsibility of a single multifunctional kinase/phosphotransferase, ITPK1. This enzyme dynamically couples the cellular levels of Ins(3,4,5,6)P₄ to the receptor-dependent hydrolysis of inositol lipids by phospholipase C. This is a biologically significant event because Ins(3,4,5,6)P₄ regulates the conductance of a specialized class of chloride ion channels, which regulate many cellular functions including epithelial salt and fluid secretion, synaptic efficacy, bone remodelling, tumor cell migration, insulin release from pancreatic β-cells, and inflammatory responses. This review assesses the current state of our knowledge of this versatile and ubiquitous signalling cascade.     

Syntheses and thermal characterization of the magnesium phthalocyanine complexes with 2-methoxy- and with 2-ethoxyethanol. X-ray structure of (2-ethoxyethanol)magnesium phthalocyanine

The syntheses of the magnesium phthalocyanine complexes with dry 2-methoxy-ethanol and 2-ethoxyethanol have been performed by recrystallization method using both anhydrous MgPc and aquated magnesium phthalocyanine, MgPcH₂O, as a starting material. It has turned out that in the temperature range below ca. 140 °C, the bi-axially ligated complexes are formed, i.e., MgPc(2-methoxyethanol)₂ and MgPc(2-ethoxyethanol)₂ with 4 + 2 coordination of Mg, whereas at higher temperatures, up to about 200 °C, the mono-axially ligated complexes are stable, i.e., MgPc(2-methoxyethanol) and MgPc(2-ethoxyethanol) with 4 + 1 coordination of Mg.The single crystal structure of MgPc(2-ethoxyethanol) complex has been determined. The central Mg atom is displaced towards the hydroxyl group of the ligand by about 0.37 Å from the (N-isoindole)₄ plane of the Pc ring. Hydrogen bonds of the type O-H?N between the hydroxyl groups of 2-ethoxyethanol and one of the azamethine N-atoms of the Pc ring link the molecules related by a centre of symmetry. Such a packing arrangement in the crystal leads to a dimerization with the ligand-to-Pc connections. The syntheses, thermogravimetric results and structure characteristics are compared with the known MgPc complexes with O- and N-donating molecules.     

Effect of heteroatom-doped carbon quantum dots on the red emission of metal-conjugated phthalocyanines through hybridization

Quantum dots (QDs) are significant fluorescent materials for energy transfer studies with phthalocyanines (Pcs) and phthalocyanine (Pc)-like biomolecules (such as chlorophylls). Carbon-based QDs, especially, have been used in numerous studies concerning energy transfer with chlorophylls, but the numbers of studies concerning energy transfer between phthalocyanines and carbon-based QDs are limited. In this study, peripherally, hydroxythioethyl terminal group substituted metal-free phthalocyanine (H₂Pc) and zinc phthalocyanine (ZnPc) were noncovalently (electrostatic and/or π–π interaction) attached to carbon QDs containing boron and nitrogen to form QD-Pc nanoconjugates. The QD-Pc conjugates were characterized using different spectroscopic techniques (Fourier transform infrared spectroscopy and transmission electron microscopy). The absorption and fluorescence properties of QD-Pc structures in solution were studied. It was found that the quantum yields of the QDs slightly decreased from 30% to 25% upon doping the QDs with heteroatoms B and N. Förster resonance energy transfer efficiency was calculated as 33% for BCN-QD/ZnPc. For the other conjugates, almost no energy transfer from QDs to Pc cores was observed. It was shown that the energy transfer between QDs to Pc cores was completely different from the energy transfer between QDs and photosynthetic pigments, and therefore we concluded that heteroatom doping in the QD structure and the existence of zinc metal in the phthalocyanine structure is obligatory for an efficient energy transfer.     

Binding Modes of Phthalocyanines to Amyloid β Peptide and Their Effects on Amyloid Fibril Formation

The inherent tendency of proteins to convert from their native states into amyloid aggregates is associated with a range of human disorders, including Alzheimer’s and Parkinson’s diseases. In that sense, the use of small molecules as probes for the structural and toxic mechanism related to amyloid aggregation has become an active area of research. Compared with other compounds, the structural and molecular basis behind the inhibitory interaction of phthalocyanine tetrasulfonate (PcTS) with proteins such as αS and tau has been well established, contributing to a better understanding of the amyloid aggregation process in these proteins. We present here the structural characterization of the binding of PcTS and its Cu(II) and Zn(II)-loaded forms to the amyloid β-peptide (Aβ) and the impact of these interactions on the peptide amyloid fibril assembly. Elucidation of the PcTS binding modes to Aβ₄₀ revealed the involvement of specific aromatic and hydrophobic interactions in the formation of the Aβ₄₀-PcTS complex, ascribed to a binding mode in which the planarity and hydrophobicity of the aromatic ring system in the phthalocyanine act as main structural determinants for the interaction. Our results demonstrated that formation of the Aβ₄₀-PcTS complex does not interfere with the progression of the peptide toward the formation of amyloid fibrils. On the other hand, conjugation of Zn(II) but not Cu(II) at the center of the PcTS macrocyclic ring modified substantially the binding profile of this phthalocyanine to Aβ₄₀ and became crucial to reverse the effects of metal-free PcTS on the fibril assembly of the peptide. Overall, our results provide a firm basis to understand the structural rules directing phthalocyanine-protein interactions and their implications on the amyloid fibril assembly of the target proteins; in particular, our results contradict the hypothesis that PcTS might have similar mechanisms of action in slowing the formation of a variety of pathological aggregates.     

Influence of potassium and manganese on growth and uptake of magnesium by soybeans (Glycine max (L.) Merr. cv. Bragg)

Summary Soybean (Glycine max (L) Merr. cv. Bragg) seedlings were grown in nutrient solutions to evaluate the response to manganese nutrition as affected by potassium supply. In solutions containing 275 M manganese, increasing the solution concentration of potassium from 1 mM to 10 mM alleviated symptoms of manganese toxicity, decreased manganese concentrations in the leaves and increased dry matter yields of the plants. The reduction in manganese toxicity was brought about by a reduced rate of root absorption of manganese at high potassium supply levels.Increasing the supply of either potassium or manganese decreased the leaf concentration of magnesium although there were no apparent symptoms of magnesium deficiency in any treatment. The reduced concentration of magnesium in the leaves was due to effects of potassium and manganese on the rate of root absorption of magnesium.Under manganese deficiency conditions, growth was reduced and manganese concentrations in plant parts were very low; there was no effect of potassium supply when manganese was absent from the nutrient solution.     

β-Carboline glucoalkaloids from Strychnos mellodora

Two new N_b-methylated β-carbolinium glucoalkaloids, 3,4,5,6-tetradehydropalicoside and 3,4,5,6-tetradehydrodolichantoside, together with the known β-carboline compounds desoxycordifoline (β-carboline 3-carboxylate glucoalkaloid) and melinonine F (N_b-methylated harmanium cation), were isolated from Strychnos mellodora stembark. The structures of the compounds were elucidated on the basis of spectroscopic studies.     

Suppression of Oxygen Evolving System in Spinach Chloroplast Membranes due to Release of Manganese on Exposure to Osmotic Stress in vitro in Presence of Magnesium

When spinach chloroplast membranes were exposed to osmotic stress in vitro, by incubation in 1.0 M sorbitol + 10 mM MgCl₂ their oxygen evolving system was suppressed. The possible reasons for such inactivation of PS II mediated oxygen evolution were examined. There were conformational changes in the chloroplast membranes, as indicated by their absorption spectra. The pattern of sensitivity to DCMU was not altered. The sensitivity of PS II to water stress remained, even after a pre-wash treatment with NaCI (which removed 18 and 24 kD proteins) but not when the thylakoids were pretreated with NH₂0H or CaCl₂ (removed manganese and 33 kD). The manganese content of thylakoid membranes was markedly reduced under osmotic stress in presence of magnesium. We suggest that exposure of chloroplasts to 1.0 M sorbitol in presence of Mg₂₊ released manganese from thylakoid membranes, thereby leading to a suppression in oxygen evolution.     

Rat osseous plate alkaline phosphatase: mechanism of action of manganese ions

Polidocanol-solubilized osseous plate alkaline phosphatase was modulated by manganese ions in a similar way as by zinc ions. For concentrations up to 1.0 nm, the enzyme was stimulated by manganese ions, showing site-site interactions (n = 2.2). However, larger concentrations (> 0.1 m) were inhibitory. Manganese ions could play the role of zinc ions stimulating the enzyme synergistically in the presence of magnesium ions (K _d = 7.2 m; V = 1005.5 U mg^–1). Manganese ions could also play the role of magnesium ions, stimulating the enzyme synergistically in the presence of zinc ions (K _d = 2.2 m; V = 1036.7 U mg^–1). However, manganese ions could not substitute for zinc and magnesium at the same time since ion assymetry is necessary for full activity of the enzyme. A steady-state kinetic model for the modulation of enzyme activity by manganese ions is proposed.     

Synthesis and antimalarial activity of metal complexes of cross-bridged tetraazamacrocyclic ligands

Using transition metals such as manganese(II), iron(II), cobalt(II), nickel(II), copper(II), and zinc(II), several new metal complexes of cross-bridged tetraazamacrocyclic chelators namely, cyclen- and cyclam-analogs with benzyl groups, were synthesized and screened for in vitro antimalarial activity against chloroquine-resistant (W2) and chloroquine-sensitive (D6) strains of Plasmodium falciparum. The metal-free chelators tested showed little or no antimalarial activity. All the metal complexes of the dibenzyl cross-bridged cyclam ligand exhibited potent antimalarial activity. The Mn²⁺ complex of this ligand was the most potent with IC₅₀s of 0.127 and 0.157 μM against the chloroquine-sensitive (D6) and chloroquine-resistant (W2) P. falciparum strains, respectively. In general, the dibenzyl hydrophobic ligands showed better anti-malarial activity compared to the activity of monobenzyl ligands, potentially because of their higher lipophilicity and thus better cell penetration ability. The higher antimalarial activity displayed by the manganese complex for the cyclam ligand in comparison to that of the cyclen, correlates with the larger pocket of cyclam compared to that of cyclen which produces a more stable complex with the Mn²⁺. Few of the Cu²⁺ and Fe²⁺ complexes also showed improvement in activity but Ni²⁺, Co²⁺ and Zn²⁺ complexes did not show any improvement in activity upon the metal-free ligands for anti-malarial development.     

Synthesis and investigation of spectral and electrochemical properties of alkyl-substituted planar binuclear phthalocyanine complexes sharing a common naphthalene ring

Hexa-tert-butyl and dodeca-n-butyl-substituted planar binuclear phthalocyanines sharing a common naphthalene ring with Mg as a central metal were synthesized with high yields and characterized by UV/Vis spectra, luminescence spectra, NMR, electrochemical, and spectroelectrochemical measurements. On the base of these complexes, the metal-free phthalocyanine ligands and the series of binuclear phthalocyanine complexes of rare earth elements (REE) were synthesized.All compounds obtained revealed an intensive near IR-absorption reaching 855 nm for trinuclear phthalocyanine. A crucial increase in NMR spectra resolution was achieved by the addition of ethylene glycol as a disaggregating agent. Spectroelectrochemical measurements during oxidation showed reversible changes of absorbance at 709 and 800 nm.     

Effects of 17β-Estradiol and Vitamin E Treatments on Blood Trace Element and Antioxidant Enzyme Levels in Ovariectomized Rats

We investigated the effect of 17β-estradiol (E₂) alone and separately vitamin E treatment on trace element status of rats following an ovariectomic operation. Forty rats were equally divided into four groups: Group 1, control, non-ovariectomized rats; Group 2, (OVX) rats, ovariectomized under general anesthesia; Group 3, (OVX+E₂) rats, the group received a 40 μg kg⁻¹ subcutan dose of E₂ per day after ovariectomy; and Group 4, (OVX + E₂ + vitamin E) rats, received the same E₂ treatment, but with an additional 100 mg kg⁻¹ intraperitoneal dose of vitamin E per day after ovariectomy. At the end of the 30-day experiment, the rats were sacrificed and their blood was collected for the measurement of zinc, copper, iron, phosphorus, selenium, magnesium, calcium, manganese, and chromium; copper–zinc superoxide dismutase (SOD); manganese-superoxide dismutase (Mn-SOD); glutathione peroxidase (Se-GSH-Px); and catalase (CAT). The levels of zinc, copper, iron, phosphorus, selenium, calcium, chromium, and manganese and activities of SOD, Mn-SOD, Se-GSH-Px, and CAT were lower in the OVX than in the control group, but magnesium level was unaffected. However, zinc, copper, iron, phosphorus, selenium, calcium, chromium, and manganese levels and SOD, Mn-SOD, Se-GSH-Px, and CAT activities were higher under separate E₂ and E₂ + vitamin E treatments. The level of magnesium in the treated-OVX groups was not different than in the OVX group. In conclusion, E₂ treatment has an ameliorating effect on the trace element status in OVX, and this effect may be enhanced with the addition of vitamin E.     

Adhesion of Human Epidermal Keratinocytes to Laminin

We have examined the mechanism by which human epidermal keratinocytes adhere to the A/B1/B2 (α₁β₁γ₁) form of laminin. Adhesion could be completely inhibited with an antibody to the β₁ integrin subunit or a combination of antibodies recognising the α₂β₂ a₃β₁ and α₆β₄ integrins. Keratinocytes adhered in the presence of magnesium and manganese ions, but calcium ions did not support adhesion and inhibited adhesion when combined with magnesium and manganese. The effects of anti-integrin antibodies (including a stimulatory antibody to the β₁ subunit) were not influenced by specific cations, with the exception that inhibition by an antibody to α₂β₁ was abrogated by the presence of manganese ions. The E3 and E8 proteolytic fragments of laminin did not support keratinocyte adhesion and heat inactivation of the E8 site in intact laminin did not reduce adhesion. Three laminin fragments that did support adhesion were P1, E4 and E1X-Nd, P1 activity being attributable at least in part to the RGD site; antibody blocking experiments suggested that adhesion to these fragments was primarily via α₁β₃. The synthetic peptide GD-6, derived from the carboxy terminus of the laminin A chain (included within E3) did support adhesion, but the significance of this observation is unclear, since a scrambled control peptide could also support adhesion. In conclusion, keratinocyte adhesion to A/B1/B2 laminin involves three integrins and multiple binding sites that are different from those defined previously.     

Tetrapyrrole utilization by Bacteroids ruminocola.

Reduced versus oxidized difference spectra of whole cells and pyridine hemochromogens of heme-requiring isolates of Bacteroides ruminicola are altered when deuteroporphyrin or mesoporphyrin replaces protoheme as a growth factor. During growth in the presence of either deuteroporphyrin or mesoporphyrin, whole cells exhibit peaks at 545 t547, 515 to 518, and 412 to 413 nm. Pyridine hemochromogen spectra confirm the presence of meso -or deuteroheme in cells grown in the presence of meso- or deuteroporphyrin. No evidence was found for the conversion of either meso- or deuteroporphyrin to protoheme. Cells grown in the presence of the manganese of magnesium chelates of protoheme form iron-containing hemes. Neither spontaneous decomposition of noniron metalloporphyrin chelates nor spontaneous formation of hemes from Fe2+ and metal-free porphyrins was detected. Protoheme-synthesizing isolates of B. ruminicola fail to use preformed metal-free porphyrins, but form both protoheme- and deuteroheme-containing cytochromes when grown in the presence of manganese deuteroheme. Versatility in tetrapyrrole utilization by B. ruminicola appears to reflect the ability of the organism to mediate the removal of nonferrous ions and to insert Fe2+ into the tetrapyrrole nucleus. The orgamism also forms functional b-type cytochromes with prosthetic groups other than protoheme.     

Mycobacterium smegmatis DinB2 misincorporates deoxyribonucleotides and ribonucleotides during templated synthesis and lesion bypass

Mycobacterium smegmatis DinB2 is the founder of a clade of Y-family DNA polymerase that is naturally adept at utilizing rNTPs or dNTPs as substrates. Here we investigate the fidelity and lesion bypass capacity of DinB2. We report that DinB2 is an unfaithful DNA and RNA polymerase with a distinctive signature for misincorporation of dNMPs, rNMPs and oxoguanine nucleotides during templated synthesis in vitro. DinB2 has a broader mutagenic spectrum with manganese than magnesium, though low ratios of manganese to magnesium suffice to switch DinB2 to its more mutagenic mode. DinB2 discrimination against incorrect dNTPs in magnesium is primarily at the level of substrate binding affinity, rather than k_pol. DinB2 can incorporate any dNMP or rNMP opposite oxo-dG in the template strand with manganese as cofactor, with a kinetic preference for synthesis of an A:oxo-dG Hoogsteen pair. With magnesium, DinB2 is adept at synthesizing A:oxo-dG or C:oxo-dG pairs. DinB2 effectively incorporates deoxyribonucleotides, but not ribonucleotides, opposite an abasic site, with kinetic preference for dATP as the substrate. We speculate that DinB2 might contribute to mycobacterial mutagenesis, oxidative stress and quiescence, and discuss the genetic challenges to linking the polymerase biochemistry to an in vivo phenotype.     

Mucosal uptake and whole-body retention of dietary manganese are not altered in beta2-microglobulin knockout mice

To further examine the interrelationships between manganese and iron absorption, the mucosal uptake, initial rate of loss, wholebody retention, and tissue distribution of an orally administered ⁵⁴Mn radiotracer were compared between normal and β₂-microglobulin knockout [β₂m(-/-)] mice. These mutant mice are commonly used as a model for the study of human hemochromatosis, a hereditary ironoverload disease. Initial uptake of ⁵⁴Mn by the intestinal mucosa, the liver, and the brain was not different between the two strains. The mutant mice had much higher concentrations of nonheme and total iron in the liver, but hepatic manganese, copper, magnesium, and zinc concentrations were similar between the two strains. In summary, the mucosal uptake and whole-body retention of manganese and tissue manganese concentrations were not altered in β₂m(-/-) mice; this suggests that normal homeostasis of manganese is not affected by the altered HFE protein-β₂m complex in these mice. Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the US Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable. The US Department of Agriculture, Agricultural Research Service, Northern Plains Area, is an equal opportunity/affirmative action employer and all agency services are available without discrimination.     

Synthesis and antimicrobial activity of copper(II) and manganese(II) α,ω-dicarboxylate complexes

Copper(II) ,-dicarboxylate complexes of general formulae, [Cu(O₂C(CH₂)_nCO₂)]·xH₂O, [Cu(O₂C(CH₂)_nCO₂) (phen)₂]·xH₂O and [Cu(O₂C(CH₂)_nCO₂)(bipy)_y]·xH₂O (n=1–8; y=1, 2; phen = 1,10-phenanthroline; bipy = 2,2-bipyridine) were synthesised. These copper complexes, some related manganese(II) complexes and the metal-free ligands were screened in vitro for their ability to inhibit the growth of Candida albicans. Metal-free 1,10-phenanthroline and all of the copper(II) and manganese(II) phenanthroline complexes were potent growth inhibitors, with only one bipyridine complex, [Cu(O₂C(CH₂)CO₂)(bipy)₂]·2H₂O, having moderate activity. The remaining substances were effectively inactive. Complexes which were active against C. albicans also proved effective against C. glabrata, C. tropicalis and C. kreusi with the manganese complexes retaining superior activity. For the phenanthroline complexes the active drug species is thought to be the dication [M(phen)₂(H₂O)_n]²⁺ (M = Cu, Mn). Escherichia coli and Staphylococcus aureus were resistant to all of the metal complexes and also to metal-free 1,10-phenanthroline. Only the copper phenanthroline complexes showed intermediate activity against Pseudomonas aeruginosa.     

RNA 5′-Triphosphatase, Nucleoside Triphosphatase, and Guanylyltransferase Activities of Baculovirus LEF-4 Protein

Autographa californica nuclear polyhedrosis virus late and very late mRNAs are transcribed by an RNA polymerase consisting of four virus-encoded polypeptides: LEF-8, LEF-9, LEF-4, and p47. The 464-amino-acid LEF-4 subunit contains the signature motifs of GTP:RNA guanylyltransferases (capping enzymes). Here, we show that the purified recombinant LEF-4 protein catalyzes two reactions involved in RNA cap formation. LEF-4 is an RNA 5′-triphosphatase that hydrolyzes the γ phosphate of triphosphate-terminated RNA and a guanylyltransferase that reacts with GTP to form a covalent protein-guanylate adduct. The RNA triphosphatase activity depends absolutely on a divalent cation; the cofactor requirement is satisfied by either magnesium or manganese. LEF-4 also hydrolyzes ATP to ADP and P_i (K_m = 43 μM ATP; V_max = 30 s⁻¹) and GTP to GDP and P_i. The LEF-4 nucleoside triphosphatase (NTPase) is activated by manganese or cobalt but not by magnesium. The RNA triphosphatase and NTPase activities of baculovirus LEF-4 resemble those of the vaccinia virus and Saccharomyces cerevisiae mRNA capping enzymes. We suggest that these proteins comprise a novel family of metal-dependent triphosphatases.     

Metal cation controls phosphate release in the myosin ATPase

Myosin is an enzyme that utilizes ATP to produce a conformational change generating a force. The kinetics of the myosin reverse recovery stroke depends on the metal cation complexed with ATP. The reverse recovery stroke is slow for MgATP and fast for MnATP. The metal ion coordinates the γ phosphate of ATP in the myosin active site. It is accepted that the reverse recovery stroke is correlated with the phosphate release; therefore, magnesium “holds” phosphate tighter than manganese. Magnesium and manganese are similar ions in terms of their chemical properties and the shell complexation; hence, we propose to use these ions to study the mechanism of the phosphate release. Analysis of octahedral complexes of magnesium and manganese show that the partial charge of magnesium is higher than that of manganese and the slightly larger size of manganese ion makes its ionic potential smaller. We hypothesize that electrostatics play a role in keeping and releasing the abstracted γ phosphate in the active site, and the stronger electric charge of magnesium ion holds γ phosphate tighter. We used stable myosin–nucleotide analog complex and Raman spectroscopy to examine the effect of the metal cation on the relative position of γ phosphate analog in the active site. We found that in the manganese complex, the γ phosphate analog is 0.01 nm further away from ADP than in the magnesium complex. We conclude that the ionic potential of the metal cation plays a role in the retention of the abstracted phosphate.