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Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010Computational Studies of the Stability and Chirality ofSelf-Assembled Complexes of a Novel Iron(II)-BindingInsulin DerivativeNiels Johan Christensen 1 , Henrik K. Munch 2 , Søren Thiis Heide 1 ,Thomas Hoeg-Jensen 3 , Peter Waaben Thulstrup 1 and Knud J. Jensen 21 Bioinorganic Chemistry, Department of Basic Sciences and Environment, Faculty ofLife Sciences, University of Copenhagen, Frederiksberg C, 1871, Denmark;2 Bioorganic Chemistry, Department of Basic Sciences and Environment, Faculty ofLife Sciences, University of Copenhagen, Frederiksberg C, 1871, Denmark;3 Novo Nordisk, Maaloev, 2760, DenmarkIntroductionIn a concurrent study, we demonstrate experimental evidence for the Fe(II) directedself-assembly of a novel insulin derivative (human insulin (B9Asp + B27Glu) [1])functionalized with bipyridine through the B29Lys side-chain or through the N-terminal ofthe A-chain (Figure 1). In the following, these engineered insulin monomers shall bereferred to as B29 and A1, where no confusion will arise. Here we present initialcomputational studies on self-assembled states involving the chelation of Fe(II) by threeB29 or A1 monomers.Results and DiscussionMeasurements confirm that the self-assembled insulin structures are trimers containing aFe(II) ion coordinated by three bipyridine ligands. This information is consistent with fourmodes of association, as illustrated with tentative in silico models in Figure 2 in the case ofB29. We have employed time-dependent densityfunctional theory (TDDFT) calculations at theRB3lYP/6-31G(d) level of theory on the coordinationenvironment around the Fe(II) ion to predict circulardichroism (CD) spectra of the four stereoisomers of theB29A1Fig. 1. Models of insulin X2monomers functionalized withbipyridine at the B29 or A1position.self-assembled insulin trimer. The TDDFT calculationswere carried out on small models truncated close to thebipyridine moiety, allowing representation of the Fe(II)coordination environment of both A1 and B29 trimers.The RB3LYP/6-31G(d) geometries and relative SCFenergies are shown in Figure 3. It is evident that themeridional arrangement is disfavored by 2.1 kJ/molwith respect to the facial arrangement for these models.With the chosen truncation, the and forms areenantiomers and thus have identical energies. This isnot the case for the actual insulin trimers, and we are currently studying more realisticmodels to elucidate / preferences. Figure 4 shows the measured CD spectra and spectrapredicted from the model systems. Although the calculated spectra are blue-shifted-fac -fac -mer -merFig. 2. Three insulin X2 monomers funtionalized with bipyridine (B29 or A1 of Figure 1)can form four types of coordination complex with Fe(II), differing in the chirality ( or )of the tris-bipyrdine complex and the facial (fac) or meridional (mer) arrangement ofsubstituents. The complexes shown are tentative in silico models corresponding toB29-bipy functionalized insulin.628