Modeling of metal interaction geometries for protein-ligand docking

The accurate modeling of metal coordination geometries plays an important role for structure-based drug design applied to metalloenzymes. For the development of a new metal interaction model, we perform a statistical analysis of metal interaction geometries that are relevant to protein-ligand complexes. A total of 43,061 metal sites of the Protein Data Bank (PDB), containing amongst others magnesium, calcium, zinc, iron, manganese, copper, cadmium, cobalt, and nickel, were evaluated according to their metal coordination geometry. Based on statistical analysis, we derived a model for the automatic calculation and definition of metal interaction geometries for the purpose of molecular docking analyses. It includes the identification of the metal-coordinating ligands, the calculation of the coordination geometry and the superposition of ideal polyhedra to identify the optimal positions for free coordination sites. The new interaction model was integrated in the docking software FlexX and evaluated on a data set of 103 metalloprotein-ligand complexes, which were extracted from the PDB. In a first step, the quality of the automatic calculation of the metal coordination geometry was analyzed. In 74% of the cases, the correct prediction of the coordination geometry could be determined on the basis of the protein structure alone. Secondly, the new metal interaction model was tested in terms of predicting protein-ligand complexes. In the majority of test cases, the new interaction model resulted in an improved docking accuracy of the top ranking placements.     

Bidentate and tridentate metal-ion coordination states within ternary complexes of RB69 DNA polymerase

Two divalent metal ions are required for primer‐extension catalyzed by DNA polymerases. One metal ion brings the 3′‐hydroxyl of the primer terminus and the α‐phosphorus atom of incoming dNTP together for bond formation so that the catalytically relevant conformation of the triphosphate tail of the dNTP is in an α,β,γ‐tridentate coordination complex with the second metal ion required for proper substrate alignment. A probable base selectivity mechanism derived from structural studies on Dpo4 suggests that the inability of mispaired dNTPs to form a substrate‐aligned, tridentate coordination complex could effectively cause the mispaired dNTPs to be rejected before catalysis. Nevertheless, we found that mispaired dNTPs can actually form a properly aligned tridentate coordination complex. However, complementary dNTPs occasionally form misaligned complexes with mutant RB69 DNA polymerases (RB69pols) that are not in a tridentate coordination state. Here, we report finding a β,γ‐bidentate coordination complex that contained the complementary dUpNpp opposite dA in the structure of a ternary complex formed by the wild type RB69pol at 1.88 Å resolution. Our observations suggest that several distinct metal‐ion coordination states can exist at the ground state in the polymerase active site and that base selectivity is unlikely to be based on metal‐ion coordination alone.     

The best-fit size and geometry of metal ions for coordination with nitrogen-donor macrocycles

The technique of calculating the strain energy of metal ion complexes as a function of metal to ligand bond length (Hancock and McDougall, J. Am. Chem. Soc., 102 (1980) 6553) is used to study best-fit sizes of metal ions for coordinating with tetraaza and triaza macrocyles. In addition to varying the metal to ligand bond length in the calculations, different coordination geometries of the metal ion are also examined. The metal to nitrogen (M-N) bond lengths, and coordination geometries, that give lowest energies for several N-donor macrocyles, are calculated by molecular mechanics, and 16-aneN₄ (1,5,9,13-tetraazacyclohexadecane) is found, contrary to popular belief, to coordinate best with very small metal ions, with lowest energy occurring for a slightly flattened tetrahedral metal ion of M-N length = 1.81 Å. The best-fit size and geometry for coordination in 12-aneN₄ (1,4,7,10-tetraazacyclotetradecane) is an M-N length of 2.15 Å and square pyramidal geometry, and with cyclam (1,4,8,11-tetraazacyclotetradecane) an M-N length of 2.06 Å and planar coordination that is approximately square.     

A tetrahedral coordination of Zinc during transmembrane transport by P-type Zn(2+)-ATPases

Zn(2+) is an essential transition metal required in trace amounts by all living organisms. However, metal excess is cytotoxic and leads to cell damage. Cells rely on transmembrane transporters, with the assistance of other proteins, to establish and maintain Zn(2+) homeostasis. Metal coordination during transport is key to specific transport and unidirectional translocation without the backward release of free metal. The coordination details of Zn(2+) at the transmembrane metal binding site responsible for transport have now been established. Escherichia coli ZntA is a well-characterized Zn(2+)-ATPase responsible for intracellular Zn(2+) efflux. A truncated form of the protein lacking regulatory metal sites and retaining the transport site was constructed. Metrical parameters of the metal-ligand coordination geometry for the zinc bound isolated form were characterized using x-ray absorption spectroscopy (XAS). Our data support a nearest neighbor ligand environment of (O/N)(2)S(2) that is compatible with the proposed invariant metal coordinating residues present in the transmembrane region. This ligand identification and the calculated bond lengths support a tetrahedral coordination geometry for Zn(2+) bound to the TM-MBS of P-type ATPase transporters.     

Probing determinants of the metal ion selectivity in carbonic anhydrase using mutagenesis

Few studies measuring thermodynamic metal ion selectivity of metalloproteins have been performed, and the major determinants of metal ion selectivity in proteins are not yet well understood. Several features of metal ion binding sites and metal coordination have been hypothesized to alter the transition metal selectivity of chelators, including (1) the polarizability of the coordinating atom, (2) the relative sizes of the binding site and the metal ion, and (3) the metal ion binding site geometry. To test these hypotheses, we have measured the metal ion affinity and selectivity of a prototypical zinc enzyme, human carbonic anhydrase II (CAII), and a number of active site variants where one of the coordinating ligands is substituted by another side chain capable of coordinating metal. CAII and almost all of the variants follow the inherent metal ion affinity trend suggested by the Irving-Williams series, demonstrating that this trend operates within proteins as well as within small molecule chelators and may be a dominant factor in metal ion selectivity in biology. Neither the polarizability of the liganding side chains nor the size of the metal ion binding site correlates strongly with metal ion specificity; instead, changes in metal ion specificity in the variants correlate with the preferred coordination number and geometry of the metal ion. This correlation suggests that a primary feature driving deviations from the inherent ligand affinity trend is the positioning of active site groups such that a given metal ion can adopt a preferred coordination number/geometry.     

Individual metal ligands play distinct functional roles in the zinc sensor Staphylococcus aureus CzrA

Recent studies on metalloregulatory proteins suggest that coordination number/geometry and metal ion availability in a host cytosol are key determinants for biological specificity. Here, we investigate the contribution that individual metal ligands of the alpha5 sensing site of Staphylococcus aureus CzrA (Asp84, His86, His97', and His100') make to in vitro metal ion binding affinity, coordination geometry, and allosteric negative regulation of DNA operator/promoter region binding. All ligand substitution mutants exhibit significantly reduced metal ion binding affinity (K(Me)) by > or =10(3) M(-1). Substitutions of Asp84 and His97 give rise to non-native coordination geometries upon metal binding and are non-functional in allosteric coupling of metal and DNA binding (DeltaG(coupling) approximately 0 kcal mol(-1)). In contrast, His86 and His100 could be readily substituted with potentially liganding (Asp, Glu) and poorly liganding (Asn, Gln) residues with significant native-like tetrahedral metal coordination geometry retained in these mutants, leading to strong functional coupling (DeltaG(coupling) > or = +3.0 kcal mol(-1)). 1H-(15)N heteronuclear single quantum coherence (HSQC) spectra of wild-type and mutant CzrAs reveal that all H86 and H100 substitution mutants undergo 4 degrees structural switching on binding Zn(II), while D84N, H97N and H97D CzrAs do not. Thus, only those variant CzrAs that retain some tetrahedral coordination geometry characteristic of wild-type CzrA upon metal binding are capable of driving 4 degrees structural conformational changes linked to allosteric regulation of DNA binding in vitro, irrespective of the magnitude of K(Me).     

A tetrahedral coordination of Zinc during transmembrane transport by P-type Zn2+-ATPases

Zn²⁺ is an essential transition metal required in trace amounts by all living organisms. However, metal excess is cytotoxic and leads to cell damage. Cells rely on transmembrane transporters, with the assistance of other proteins, to establish and maintain Zn²⁺ homeostasis. Metal coordination during transport is key to specific transport and unidirectional translocation without the backward release of free metal. The coordination details of Zn²⁺ at the transmembrane metal binding site responsible for transport have now been established. Escherichia coli ZntA is a well-characterized Zn²⁺-ATPase responsible for intracellular Zn²⁺ efflux. A truncated form of the protein lacking regulatory metal sites and retaining the transport site was constructed. Metrical parameters of the metal–ligand coordination geometry for the zinc bound isolated form were characterized using x-ray absorption spectroscopy (XAS). Our data support a nearest neighbor ligand environment of (O/N)₂S₂ that is compatible with the proposed invariant metal coordinating residues present in the transmembrane region. This ligand identification and the calculated bond lengths support a tetrahedral coordination geometry for Zn²⁺ bound to the TM-MBS of P-type ATPase transporters.     

From cisplatin to artificial nucleases — the role of metal ion-nucleic acid interactions in biology

Metal ions and metal coordination compounds bind to nucleic acids in a variety of ways, ranging from weak electrostatic interactions via hydrogen bonding and/or van der Waals forces to strong covalent binding. Metal ions naturally take part in the formation and the degradation of nucleic acids, and the propensity of certain metal coordination compounds to bind to nucleic acids, notably DNA, is enploited in cancer chemotherapy. Moreover, metal compounds have a wide potential as chemical probes for nucleic acid structures and as tools for nucleic acid processing.     

Syntheses and structures of M(L)(X)BPh4 complexes {M=Co(II), Zn(II); L=cis-1,3,5-tris[3-(2-furyl)prop-2-enylideneamino]cyclohexane, X=OAc, NO3}: structural models of the active site of carbonic anhydrase

The metal coordination geometries in the structures of the zinc(II) and cobalt(II) complexes of the ligand cis-1,3,5-tris[3-(2-furyl)prop-2-enylideneamino]cyclohexane (fr-protach) and with the anions nitrate and acetate are structural models for the active site of carbonic anhydrase. The acetate structures show a striking structural correlation with the metal coordination environments in the known bicarbonate forms of the enzyme. Such structures provide a basis for understanding the marked effect of different metal substitution on the catalytic rate of the enzyme.     

Complexes of amylose and amylopectins with multivalent metal salts

Metal cations [Cu(II), Fe(III), Mn(II), and Ni(II)] are ligated by amylose as well as potato, and corn amylopectins as proven by electron paramagnetic resonance spectra and conductivity measurements. The hydroxyl groups of polysaccharides are the coordination sites. Isolated starch polysaccharides did not coordinate to metal ions so well as starch did. The resulting polycenter Werner complexes were mainly square planar species. The ligation of the central metal atoms resulted in a variation of the thermal stability, pathway, and rate of thermal decomposition of starch as proven by thermogravimetric (TG, DTG) and scanning differential calorimetric measurements. Frequently, amylose and potato amylopectin willingly formed clathrates in which the water molecules were caged. The mode of the coordination of the hydroxyl groups to the central metal atom controlled the clathrate formation from amylose and in the case of potato amylopectin metal atoms bound to the phosphoric acid moiety formed cage by coordination of the hydroxyl groups to them. Coordination to selected metal salts controls pathway and products of polysaccharide ligand thermolysis.     

Designed inhibitors of horse liver alcohol dehydrogenase. Effect of active-site metal ion substitution

3-(p-Butoxyphenyl)propionamide, -thioamide and -hydrazide and the formamide of p-butoxybenzylamine were tested as inhibitors of cadmium(II) and cobalt(II) active-site substituted alcohol dehydrogenase. The results agree with a direct coordination of these inhibitors except for the hydrazide to the active-site metal ion, in the enzyme-NADH-inhibitor complex. The hydrazide might be situated at some distance from the metal ion without a direct coordination bond.     

Hydrogen binding in coordination compounds of 3(5)-(4-methoxyphenyl)pyrazole

The solid state structures of 3(5)-(4-methoxyphenyl)pyrazole and its coordination compounds with a series of two valent transition metals have been investigated. Since pyrazoles provide not only a nitrogen donor site for the coordination to metal ions, but also an additional N–H function, they are ideal ligands for the formation of hydrogen bound coordination polymers or for the implementation of secondary interactions with other ligands bound to the same central ion, resulting in a rigid ligand environment at the central metal. We chose cobalt, nickel, palladium, copper and zinc as twofold positively charged Lewis acids preferring coordination numbers of four and six to prove the capability of pyrazole to undergo intramolecular hydrogen bonds. In the four-coordinate mode, either tetrahedral (Zn²⁺) or square planar coordination geometries (Pd²⁺) are possible, providing different geometric restrictions for hydrogen bonding.     

Structural influence of hydrophobic core residues on metal binding and specificity in carbonic anhydrase II

Aromatic residues in the hydrophobic core of human carbonic anhydrase II (CAII) influence metal ion binding in the active site. Residues F93, F95, and W97 are contained in a beta-strand that also contains two zinc ligands, H94 and H96. The aromatic amino acids contribute to the high zinc affinity and slow zinc dissociation rate constant of CAII [Hunt, J. A., and Fierke, C. A. (1997) J. Biol. Chem. 272, 20364-20372]. Substitution of these aromatic amino acids with smaller side chains enhances Cu(2+) affinity while decreasing Co(2+) and Zn(2+) affinity [Hunt, J. A., Mahiuddin, A., & Fierke, C. A. (1999) Biochemistry 38, 9054-9062]. Here, X-ray crystal structures of zinc-bound F93I/F95M/W97V and F93S/F95L/W97M CAIIs reveal the introduction of new cavities in the hydrophobic core, compensatory movements of surrounding side chains, and the incorporation of buried water molecules; nevertheless, the enzyme maintains tetrahedral zinc coordination geometry. However, a conformational change of direct metal ligand H94 as well as indirect (i.e., "second-shell") ligand Q92 accompanies metal release in both F93I/F95M/W97V and F93S/F95L/W97M CAIIs, thereby eliminating preorientation of the histidine ligands with tetrahedral geometry in the apoenzyme. Only one cobalt-bound variant, F93I/F95M/W97V CAII, maintains tetrahedral metal coordination geometry; F93S/F95L/W97M CAII binds Co(2+) with trigonal bipyramidal coordination geometry due to the addition of azide anion to the metal coordination polyhedron. The copper-bound variants exhibit either square pyramidal or trigonal bipyramidal metal coordination geometry due to the addition of a second solvent molecule to the metal coordination polyhedron. The key finding of this work is that aromatic core residues serve as anchors that help to preorient direct and second-shell ligands to optimize zinc binding geometry and destabilize alternative geometries. These geometrical constraints are likely a main determinant of the enhanced zinc/copper specificity of CAII as compared to small molecule chelators.     

Substrate and metal ion promiscuity in mannosylglycerate synthase

The enzymatic transfer of the sugar mannose from activated sugar donors is central to the synthesis of a wide range of biologically significant polysaccharides and glycoconjugates. In addition to their importance in cellular biology, mannosyltransferases also provide model systems with which to study catalytic mechanisms of glycosyl transfer. Mannosylglycerate synthase (MGS) catalyzes the synthesis of α-mannosyl-D-glycerate using GDP-mannose as the preferred donor species, a reaction that occurs with a net retention of anomeric configuration. Past work has shown that the Rhodothermus marinus MGS, classified as a GT78 glycosyltransferase, displays a GT-A fold and performs catalysis in a metal ion-dependent manner. MGS shows very unusual metal ion dependences with Mg(2+) and Ca(2+) and, to a lesser extent, Mn(2+), Ni(2+), and Co(2+), thus facilitating catalysis. Here, we probe these dependences through kinetic and calorimetric analyses of wild-type and site-directed variants of the enzyme. Mutation of residues that interact with the guanine base of GDP are correlated with a higher k(cat) value, whereas substitution of His-217, a key component of the metal coordination site, results in a change in metal specificity to Mn(2+). Structural analyses of MGS complexes not only provide insight into metal coordination but also how lactate can function as an alternative acceptor to glycerate. These studies highlight the role of flexible loops in the active center and the subsequent coordination of the divalent metal ion as key factors in MGS catalysis and metal ion dependence. Furthermore, Tyr-220, located on a flexible loop whose conformation is likely influenced by metal binding, also plays a critical role in substrate binding.     

Ab Initio Coordination Chemistry for Nickel Chelation Motifs

Chelation therapy is one of the most appreciated methods in the treatment of metal induced disease predisposition. Coordination chemistry provides a way to understand metal association in biological structures. In this work we have implemented coordination chemistry to study nickel coordination due to its high impact in industrial usage and thereby health consequences. This paper reports the analysis of nickel coordination from a large dataset of nickel bound structures and sequences. Coordination patterns predicted from the structures are reported in terms of donors, chelate length, coordination number, chelate geometry, structural fold and architecture. The analysis revealed histidine as the most favored residue in nickel coordination. The most common chelates identified were histidine based namely HHH, HDH, HEH and HH spaced at specific intervals. Though a maximum coordination number of 8 was observed, the presence of a single protein donor was noted to be mandatory in nickel coordination. The coordination pattern did not reveal any specific fold, nevertheless we report preferable residue spacing for specific structural architecture. In contrast, the analysis of nickel binding proteins from bacterial and archeal species revealed no common coordination patterns. Nickel binding sequence motifs were noted to be organism specific and protein class specific. As a result we identified about 13 signatures derived from 13 classes of nickel binding proteins. The specifications on nickel coordination presented in this paper will prove beneficial for developing better chelation strategies.     

Metal induced structural changes observed in hexameric insulin

The metal ions in insulin hexamer play a crucial role in the T to R conformational transitions. We have determined the crystal structures of 2Mn2+, 1Rb1+ and 4Ni2+ human arg-insulin and compared them with the 2Zn2+ structure. The first two structures exist in the T3R3f state like the native 2Zn2+ arg-insulin, while the 4Ni2+ adopts a T6 conformation. The metal coordination is found to be tetrahedral in all the structures except that of nickel where a dual octahedral and tetrahedral coordination is found at one site. Rubidium occupies only one of the high affinity metal binding sites. The metal induced structural changes observed, have been explained.     

Complexes of dibenzylsulfoxide with the first series transition metal nitrates

Under carefully controlled experimental conditions, a series of metal nitrate complexes of dibenzylsulfoxide (DBSO) have been synthesized and isolated (for the dipositive Co, Ni, Cu and Zn elements). The compounds were characterized by use of infrared-, ultraviolet-, and visible spectra. Their stoichiometric compositions appeared to be [M- (DBSO)₃(NO₃)₂], where MCo, Ni and Cu; and [M(DBSO)_3.5(NO₃)₂], where MZn. The additional information concerning the nature of bonding and structural geometry was derived from X-ray diffraction analysis, molecular conductivities, molecular weight and magnetic susceptibility measurements. The infrared spectra indicated that, in all cases, coordination occurred through the oxygen atom of the DBSO ligand. Both analyses and spectral studies suggest that the transition metal ions manifest a coordination number of six. The slight decrease in metal ion radius across the first transition metal series does not affect the number of DBSO molecules bonded to the M²⁺ ion, but the only variation in total coordination of cation might be attributed to the DBSO competition with nitrato groups for the coordination site due to the ability of the anion to enter the coordination sphere.