Zinc(II) Complexes of Tripodal Peptides Mimicking the Zinc(II)-Coordination Structure of Carbonic Anhydrase Ulrike Herr, a Werner Spahl, a Gunter Trojandt, a Wolfgang Steglich, a, * Florian Thaler b and Rudi van Eldik b a Institut fu Èr Organische Chemie, Universita Èt Mu Ènchen, Karlstraûe 23, D-80333 Mu Ènchen, Germany b Institut fu Èr Anorganische Chemie, Universita Èt Erlangen-Nu Èrnberg, Egerlandstraûe 1, D-91058 Erlangen, Germany Received 14 April 1998; accepted 13 July 1998 AbstractÐTwo new tripodal peptide ligands with histidine side chains have been synthesized and were shown to form stable zinc(II) complexes. Their NMR and mass spectra indicate a structure that is analogous to the active center of carbonic anhydrase. Both the ligands and the zinc complexes were titrated potentiometrically in order to obtain the pK a values for the coordinated water of the zinc complexes; due to the low solubility of the complexes only estimates could be obtained. # 1999 Elsevier Science Ltd. All rights reserved. Introduction The enzyme carbonic anhydrase (CAII) catalyzes the reversible hydration of carbon dioxide with an impress- ively high turnover number (10 6 s 1 , eq (1)). 1,2 CO 2  H 2 O  HCO  3  H  1 This crucial and rather simple reaction has fascinated numerous research groups. Extensive studies have been carried out to elucidate the chemical and structural aspects of its mechanism. 2±8 The results of these inves- tigations help to shed light upon the evolution of CAII into biochemically important noncatalytical functions, such as signal transduction. 9±13 The structures of CAII in the zinc±water, zinc±hydroxide, and zinc±bicarbonate forms that are involved in the reaction, are well characterized. 2,5,14,15 The active site of CAII consists of a zinc(II) ion pseudotetrahedrally coordinated by three histidine imidazoles (His-96, -94, and -119) and either a water molecule or an OH  ion in the fourth position as depicted in Figure 1. 14 The zinc(II)-bound water molecule is deprotonated to form a zinc(II)-bound hydroxide species, which is widely accepted to be the catalytically active com- plex. 13,16,17 Thus, the ability of CAII to hydrate CO 2 is characterized by a pK a value (7) of this deprotonation. 18 To mimic the zinc(II)-coordination structure or the function of the zinc(II) ion at the active site, several mononuclear model zinc(II) complexes have been designed and investigated (Fig. 2). 19±37 Up to now, the best functional models of CAII are the zinc(II) complexes of 1,5,9-triazacyclododecane ([12]aneN 3 ) (1) 24±27 and 1,4,7,10-tetraazacyclododecane([12]aneN 4 ) (2) 28±30 in the forms ZnL(H 2 O) 2+ and ZnL(OH  ) 2+ . Woolley's enzyme model 3 31 was also successful in reproducing catalysis of carbon dioxide hydrolysis. The pK a values of complexes 1a±3a were reported to approximate that of CAII very closely. The Zn(II)±OH  complex 4 32±34 was the ®rst structural model of the active site of CAII to be isolated. Recently, Parkin et al. 37 synthesized and characterized the mononuclear zinc(II)-complex 5, in which the zinc(II) ion is coordi- nated by three imidazole moieties. However, there have been relatively few attempts to design and prepare zinc(II)-binding peptides. 38 Here, we report the synthesis of two peptide ligands with three histidine side chains featuring an environment analo- gous to the active center of CAII. Results and Discussion Synthesis of the tripodal histidine ligands C 3 -Symmetrical molecules are of particular interest as model compounds for natural ionophores and as ligands for metal complexation. 39,40 The optically pure C 3 -symmetrical triacid (SSS)-N(BzGly*ValOH) 3 (6) 0968-0896/99/$ - see front matter # 1999 Elsevier Science Ltd. All rights reserved. PII: S0968-0896(98)00180-1 Bioorganic & Medicinal Chemistry 7 (1999) 699±707 Key words: Peptides; mimetics; histidine ligand; carbonic anhydrase; metal complex. *Corresponding author. Tel.: +49-89-5902-528; fax: +49-89-5902-604; e-mail: wos@org.chemie.uni-muenchen.de