Journal of Inorganic Biochemistry 115 (2012) 182–185

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Journal of Inorganic Biochemistry

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Plasticity in the copper–thioether bond: Manifestation in blue Cu proteins and insynthetic analogs☆

Jan Reedijk ⁎Leiden Institute of Chemistry, Leiden University, P.O. Box 9502, 2300 RA, Leiden, The NetherlandsDepartment of Chemistry, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia

☆ Paper dedicated to the memory of Hans Freeman.⁎ Leiden Institute if Chemistry, Leiden University, P.

The Netherlands.

0162-0134/$ – see front matter © 2012 Elsevier Inc. Alldoi:10.1016/j.jinorgbio.2012.01.006

a b s t r a c t

a r t i c l e i n f o

Article history:Received 27 November 2011Received in revised form 13 January 2012Accepted 13 January 2012Available online 23 January 2012

Keywords:CopperThioetherBiomimeticPlasticityZinc

The binding of thioethers to transition metals in biological and biomimetic systems is reviewed with a focuson copper. Literature data show that copper(I) ions have a stronger tendency to bind thioethers, e.g.methionine-like ligands, than the isoelectronic Zn(II) ions. The plasticity in the Cu(II) coordination sphere,and the diffuseness of the lone pair electrons of a thioether sulfur, allow Cu(II)–S(thioether) bond distancesto vary from 2.4 to 3.2 Å, as shown by an in-depth analysis of protein structures (Protein Structure Database,PDB) and molecular structures of copper coordination compounds (Cambridge Structural Database, CSD).

© 2012 Elsevier Inc. All rights reserved.

1. Introduction

When Hans Freeman reported for the first time at a Gordon Re-search Conference in 1978 and in Nature [1], on the 3D structure ofplastocyanin, he also mentioned a relatively long Cu(II)–S(thioether)bond of some 2.9 Å. His report was critically received and some of theconference attendants had difficulties to accept, by intuition only,whether such a long Cu–S bond could be relevant in proteins. Initiallythe deep blue color was thought to be due to the entatic state and ageometry in between tetrahedral and square planer, as nicelyreviewed by Martell [2] and Gray [3]. The later papers of Freeman[4–6], and that of many others in the field, for a detailed review see:[3], have shown that the Cu–S bond is present in many cases, butnot in all, and that the blue color is hardly related, if at all, by thisCu–thioether bond. The deep blue color is, as we know now, causedby the Charge Transfer (CT) in the Cu–thiolate bond [3]. The redox po-tential, however, appears to bemore related to the Cu–S(thioether) dis-tances as was recently presented [7]. For a detailed discussion of thespectroscopy and redox potential, I refer to a recent review [8]. Alsothe use of Ni(II) and Co(II) substituted at the Cu site in the blue proteinshas been providing useful spectroscopic information [9–11].

Since that ground-breaking paper of Freeman [1], it has been realizedthat thioether–metal bonds occur rather regularly in natural systems, notonly in the blue copper proteins (over 150 hits in the Protein Database(PDB), October 2011, including genetically engineered proteins), but

O. Box 9502, 2300 RA, Leiden,

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also in enzymes [12], and in copper transport systems [13] and also inmany metal coordination bonds. In Fig. 1 a general schematic structureof most of the blue sites is represented.

The lone pairs on the thioether sulfur are rather diffuse and there-fore flexible, and even a drug like cisplatin is now known to use thewell-known Cu-transport site (CTR1) to enter the cell [14,15] by re-versible Pt–methionine side-group binding. We had shown someyears before [16,17] that Pt–methionine binding can indeed be re-versible, and that these bonds may be relevant for the mechanismof action of cisplatin. Also the CueO site in multicopper oxidases is arelevant Cu transport site [18].

In this brief account I want to address the significant variation inCu–S(thioether) bond lengths, and the fact that in the case of Zn(II),which is isoelectronic with Cu(I), binding to thioethers does notoccur in natural systems at all. Given the limited space, it was impos-sible to write a comprehensive review, and I apologize that many ex-amples come from our own earlier work and that a number of otherimportant references could not be included.

2. Cu–S(thioether) bond lengths in proteins

2.1. Cu(II)

The most extensive compilation of Cu(II)–methionine bondlengths in a paper is available from the review of Gray et al. [3], forboth pseudo-tetrahedral coordination geometries (Cu–S=2.60–3.00 Å) and for trigonal bipyramidal structures (Cu–S=3.01–3.26 Å); in this last case the fifth ligand is a carbonyl oxygen (Cu–O=2.51–3.14 Å). Only in stellacyanin and in some genetically

N

N

N

N

S

S

(CH2)n

R

R

Cu

X

Y

N

N

N

N

S

R

R

Cu

X

Yn(H2C)

(CH2)n

Fig. 2. Schematic structures of a selection of coordination geometries found for Cu(II)–thioether compounds with axial and equatorial binding. R=H, Me; n=0–2; X,Y=solvent,water, anion. The benzimidazoles can be replaced by (substituted) pyrazoles, pyridines andimidazoles, leading to quite similar structures. In the right hand figure, only one thioether Sis present and 5-coordinate structures are found in such cases [30,38].

N

HN

N

HN

Cu

SR

S

R

X

Met

R

R

His

His

Cys

Fig. 1. Coordination geometries for Cu(II) in many blue-copper proteins. The unpairedelectron of Cu(II) is in the magnetic orbital, and largely overlaps with the 3 equatorialbonds to histidine (2×) and cysteine. In some azurins an additional X ligand is present(O donor from carbonyl group) at a distance of 2.8–3.2 Å; in stellacyanin themethionine isabsent [3]. In the Cu(I) forms of the proteins (not blue) the coordination geometries arehardly changed.

183J. Reedijk / Journal of Inorganic Biochemistry 115 (2012) 182–185

modified azurins the Cu(II)–thioether bond is absent, as well as in lac-case and ceruloplasmin. In all cases the remaining three ligands aretightly bound and consist of two histidines and one cysteine in an al-most planar trigonal coordination geometry. Only small variations inthese distances occur, often close to the experimental uncertainties.The intrinsic property of the d9 Cu(II) ion is its plasticity originatingfrom the Jahn–Teller effect, allowing large variations in the axial di-rections, both for trigonal-based and tetragonal-based structure[19–21]. In a more recent study on nitrate reductase with a side-oncoordinated NO ligand, Tocheva et al. reported a quite short Cu(II)–S(thioether) distance of only 2.40 Å [22]. Another very short Cu–S(met) bond in a protein is known for the Cu(B) site in a dioxygen ad-duct of peptidylglycine-α-hydroxylating monooxygenase [12], whereit is only 2.4 Å. This site is not a real blue-Cu site, however, as it has nocysteine. In this case also a bond is involved having an orientation to-wards an orbital where the unpaired electron of Cu(II) is located.

2.2. Cu(I)

The number of protein structures with rather accurate Cu(I)–S(thioether) distances is still relatively small, as in most cases theCu(II) form of the protein has been crystallized. The Cu–S distancesvary from 2.7 to 3.1 Å [3]. In the case of the Zn(II) substituted azurin,the Zn–S distance was reported to be as large as 3.40 Å, (data not inPDB [10]), suggesting at most a weak Zn–S interaction. It is remark-able that as far as known today, no zinc proteins have been reportedto contain a methionine side group as a ligand (vide infra).

More recently, it has become clear that especially in brain chemis-try, the reversible Cu–methionine bond is occurring relatively fre-quently [23–25], and the Cu–methionine bond has been used totransport a Gd compound through the blood–brain barrier [26]. How-ever, accurate Cu–S distances and detailed coordination geometriesremain still relatively scarce.

3. The Cu–S(thioether) bond lengths in biomimetic compounds

Soon after the discovery of the relevance of Cu–thioether bonds inblue copper proteins, coordination chemists [27–36], including ourlaboratory, started to design compounds where the thioether groupwas allowed to bind to Cu(II). For easy synthesis, initiallybenzimidazole-based ligands were used, later followed by syntheti-cally less-easily accessible imidazole [35,37]. Also pyrazoles havebeen used [34,35]. Over the years this work has resulted in a largenumber of compounds with a wide range of Cu–S bond lengths. A re-cent search of the CSD (Cambridge Structural Database; release No-vember 2011) gives a broad band width of observed Cu–S(thioether) distances, for both Cu(I) and Cu(II), i.e. 2.4–3.2 Å, witha peak at around 2.8 Å. The actual distance appears to depend onthe other ligands present in the compound, and – for Cu(II) –whether

or not the thioether is binding in a direction where the Cu(II) orbitalwith the unpaired electron is. When the Cu–S bond is coinciding withthe magnetic orbital of Cu(II) the bonds are usually much shorter,compared to compounds where the thioether is an axial ligand.Some special cases are schematically redrawn in Fig. 2, to illustratethe different binding possibilities. For an early review I refer to ref.[35].

In most of the cases, the thioether to Cu binding was found to bestabilized by the chelate effect. Monodentate thioether–Cu bindingis rare, but cases are known [39], especially for Cu(I). Even bridgingthioethers between 2 Cu atoms are known [40,41]. In those casesthe Cu–S can be as short as 2.22 Å [42], but also much longer in a pyr-azole case: 2.64–2.68 Å [40]. In most of such cases it involves Cu(I)compounds [42,43]. In case of an organometallic compound [44], avery short Cu–S distance of 2.17 Å has been reported. When a shortCu(II)–thioether is found, i.e. below 2.5 Å, the bond orientation is al-ways almost coinciding/overlapping with the magnetic orbital ofCu(II). Several examples have been reported, but only a selectionwill be mentioned here [27,28,32,45,46].

When the contact distance to Cu(II) is above 2.6 Å, the thioether islocated in the axial position and binds only weakly. Depending on the(flexibility of the) chelating ligand, one S can be axial, and one equa-torial, resulting in one short (b2.5) and one long bond (>2.7). How-ever, also in-between distances are known, again illustrating theflexibility and plasticity of the Cu(II) coordination.

In cases where such Cu(II) compounds are reduced to Cu(I), the 5-and 6-coordination will change to linear, with just 2 short Cu–Nbonds, and in addition 2 remote Cu–S(thioether) contacts that mayor may not be considered as bonds [31,47].

4. The Zn–S(thioether) bond lengths in coordination compounds

To understand the importance of the methionine–Cu bonds, coor-dination to Co, Ni and Zn has often been studied, including studies inthe 1980s from my lab [48] and others [46]. The structures werefound to be the same as those of the corresponding Cu(II) com-pounds, with just variations in the M–S bond distances. In the caseof Co(II), in one case spectroscopic evidence was found for non-coordination of the thioethers [48], leaving a tetrahedral cobalt(II).With Zn, we failed to prove the structures by XRD; however, recentlyKaim et al. [49] could prove the Zn–thioether bond for bis(benzimida-zole)–dithiaalkane ligands. Indeed octahedral Zn(II) with Zn–S dis-tances 2.59–2.63 Å were found; with a longer alkane bridge, a

184 J. Reedijk / Journal of Inorganic Biochemistry 115 (2012) 182–185

dinuclear compound was found, but still with Zn–S coordination(2.59–2.67 Å) [49].

Though Nature has made use of the d10 ion Cu(I) to bindthioethers from methionine, this has not happened for Zn(II), whichis isoelectronic and has a higher ionic charge. However, coordinationcompounds of Zn(II) that do contain a thioether-to-Zn bond areknown; their occurrence is small though and when we restrict our-selves to heterocyclic ligands, like imidazole, pyrazole and pyridineas mimics of histidine, there are just a few available in the CambridgeDatabase. Cases where the thioether is part of a macrocycle or con-nected to aliphatic amines will not be considered here. A few repre-sentative examples of open chain chelating ligands with connectedheterocyclics are discussed below.

An early example of 5-coordinate Zn involves the work of Teixidoret al. [50] with long Zn–S contacts of 2.63–2.74 Å, whereas Garner etal. [51] later reported a similar case, now with just one Zn–S bond(2.49 Å). With pyrazoles, Bouwman found distances of 2.67–2.68 Å[52], whereas Berreau reported much shorter distances of 2.29–2.43 Å[53]. A very long contact of over 2.80 Å was reported by Butcher et al.[54]. In almost all cases the coordination geometries are in between tri-gonal bipyramidal and square pyramidal.

For 6-coordinate Zn, examples include the above-mentioned workof Kaim et al. [49] using benzimidazole-based ligands with Zn–S dis-tances of 2.59–2.67 Å. Other interesting cases include the work ofButcher et al. [55,56] where Zn–S distances of 2.50 and 2.60 Å werefound.

5. Bioinorganic applications of compounds with a Cu–thioetherbond

The biological relevance of the Cu–thioether bond is not restrictedto blue copper proteins. The Cu–S(thioether) interaction has alsobeen applied as a drug, and as a transport agent. The oldest knowncase is the use of Tagamet (Cimetidine): 2-cyano-1-methyl- 3-(2-[(5-methyl-1H-imidazol-4-yl)methylthio]ethyl)guanidine [57]. Thisis a commonly used drug for peptic ulcer treatment. It acts as a hista-mine H2-receptor antagonist that inhibits the production of acid inthe stomach. The structure of its Cu compound [57], a compound be-lieved to be part of the mechanism, is given in Fig. 3.

More recently, it was realized that cisplatin may enter (tumor)cells using the so-called CTR-1 gate [14,15], which is the normalway of Cu to enter cells, and which has many methionines, believedto guide the Cu through. Pt compounds also have a good reversiblebinding to thioethers, as we had already proposed [16,17] beforethe CTR-1 option was published [14,15,60].

In the last few years Cu–thioether bonds have been used to bringGd-imaging reagents (MRI agents) into the brain, thereby passingthe blood–brain barrier [26]. The complicated Gd chelates have sidearms with thioether groups, like in Cimetidine, that are supposed tochelate Cu(II) and bring the Gd species into the cell.

N NH

SN

NH

HN

N

NHN

SN

HN

NH

N

Cu

Fig. 3. Structure of part of the polymeric Cu–Cimetidine nitrate compound (axial nitrileligands - from a neighbouring molecule - for Cu have been omitted: Cu…N=2.56 Å).The equatorial distances are: Cu–N=1.92 Å and Cu–S=2.42 Å [58,59].

6. Concluding remarks

In the above presented brief review of the Cu–methionine bond, Ihave tried to make clear that Nature has made a clever use of it. Onone hand, the bond is relative weak and kinetically labile for bothCu(I) and Cu(II). On the other hand, the plasticity of the Cu(II) coor-dination sphere has allowed a wide range of bond distances, both innatural systems and in biomimetic coordination compounds. The ob-servation has beenmade that Zn(II), which is isoelectronic with Cu(I),has a much smaller affinity for the thioether ligand, despite the +2charge on Zn. Nature has not used the Zn–thioether bonds, whereasin coordination chemistry, using properly chosen chelating ligands,this bond can be generated.

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