The high affinity of highly charged polynuclear platinum complexes for glycans such as heparan sulfate results in modulation of the biomolecule signaling functions leading to inhibition of angiogenesis.
Cleavage of heparan sulfate proteoglycans (HSPGs) by the enzyme heparanase modulates tumour-related events including angiogenesis, cell invasion, and metastasis. Metalloshielding of heparan sulfate (HS) by positively charged polynuclear platinum complexes (PPCs) effectively inhibits physiologically critical HS functions. Studies using bacterial P. heparinus heparinase II showed that a library of Pt complexes varying in charge and nuclearity and the presence or absence of a dangling amine inhibits the cleavage activity of the enzyme on the synthetic pentasaccharide, Fondaparinux (FPX). Charge-dependent affinity of PPC for FPX was seen in competition assays with methylene blue and ethidium bromide. The dissociation constant (K ) of TriplatinNC for FPX was directly measured by isothermal titration calorimetry (ITC). The trend in DFT calculated interaction energies with heparin fragments is consistent with the spectroscopic studies. Competitive inhibition of TAMRA-R internalization in human carcinoma (HCT116) cells along with studies in HCT116, wildtype CHO and mutant CHO-pgsA745 (lacking HS/CS) cells confirm that HSPG-mediated interactions play an important role in the cellular accumulation of PPCs.
Zn(2+) inhibits the action of several of the caspases and thus may act as a regulator of apoptosis. Reversal of this inhibition is one possible approach for the development of apoptosis-based therapies. Few studies describe the molecular details of the Zn(2+)-caspase interaction, the understanding of which is essential for the success of any therapeutic strategies. Enzyme kinetics and biophysical studies have shown that the inhibition is of mixed type with prominent (ca. 60 % of inhibition) uncompetitive characteristics and an IC50 of 0.8 μM under the conditions used. Fluorescence-based techniques confirmed that, during inhibition in the sub-micromolar range, substrate binding remains unaffected. A new zinc binding site composed of the catalytic histidine and a nearby methionine residue, rather than the catalytic histidine and cysteine dyad, is proposed based on the experimental observations. DFT models were used to demonstrate that the proposed site could be the preferred inhibitory zinc binding site.
The functional role assumed by zinc in proteins is closely tied to the variable dynamics around its coordination sphere arising by virtue of its flexibility in bonding. Modern experimental and computational methods allow the detection and study of previously unknown features of bonding between zinc and its ligands in protein environment. These discoveries are occurring just in time as novel biological functions of zinc, which involve rather unconventional coordination trends, are emerging. In this sense coordination sphere expansion of structural zinc sites, as observed in our previous experiments, is a novel phenomenon. Here we explore the electronic and structural requirements by simulating this phenomenon in structural zinc sites using DFT computations. For this purpose, we have chosen MPW1PW91 and a mixed basis set combination as the DFT method through benchmarking, because it accurately reproduces structural parameters of experimentally characterized zinc compounds. Using appropriate models, we show that the greater ionic character of zinc coordination would allow for coordination sphere expansion if the steric and electrostatic repulsions of the ligands are attenuated properly. Importantly, through the study of electronic and structural aspects of the models used, we arrive at a comprehensive bonding model, explaining the factors that influence coordination of zinc in proteins. The proposed model along with the existing knowledge would enhance our ability to predict zinc binding sites in proteins, which is today of growing importance given the predicted enormity of the zinc proteome.
Zn 2+ inhibits the action of several of the caspases and thus may act as a regulator of apoptosis. Reversal of this inhibition is one possible approach for the development of apoptosis-based therapies. Few studies describe the molecular details of the Zn 2+ -caspase interaction, the understanding of which is essential for the success of any therapeutic strategies. Enzyme kinetics and biophysical studies have shown that the inhibition is of mixed type with prominent (ca. 60 % of inhibition) uncompetitive characteristics and an IC 50 of 0.8 mm under the conditions used. Fluorescence-based techniques confirmed that, during inhibition in the sub-micromolar range, substrate binding remains unaffected. A new zinc binding site composed of the catalytic histidine and a nearby methionine residue, rather than the catalytic histidine and cysteine dyad, is proposed based on the experimental observations. DFT models were used to demonstrate that the proposed site could be the preferred inhibitory zinc binding site.
Building from tryptophan to the tryptophan-containing HIV Nucleocapsid 7 (HIVNCp7) protein we combine biophysical and computational studies to enhance stacking interactions of purines through platination. The incorporation into a weak Lewis acid electrophile, [Pt(dien)(Nucleobase)]2+ may lead to disruption of the HIVNCp7-RNA interaction.
In an approach to design drugs with higher affinity for π-π stacking and electrostatic interactions with targeted biomolecules, complexes of the type [{cis-Pt(A)2(L)}2-μ-{trans-1,4-dach}](NO3)4 ((A)2 = (NH3)2 or ethylenediamine (en), L = quinoline (quin) or benzothiazole (bztz), dach = trans-1,4-diaminocyclohexane) were synthesized. The quinoline complex, [{cis-Pt(en)(quin)}2-μ-(dach)](NO3)4 (9) was synthesized from the precursor K[PtCl3(quin)] (1) while the benzothiazole complexes, [{cis-Pt(A)2(bztz)}2-μ-(dach)](NO3)4 ((A)2 = (NH3)2 (10); (A)2 = en (11)) were synthesized from the precursors cis-[Pt(A)2Cl(bztz)] ((A)2 = (NH3)2 (7); (A)2 = en (8)). Their interactions with N-acetyltryptophan and a model pentapeptide (N-AcWLASW-OH) modeled on the pentapeptide recognition sequence of p53-mdm2 (FLASW) were examined by fluorescence spectroscopy. The dinuclear complexes were found to be significantly stronger quenchers of fluorescence than their mononuclear Pt analogs. Molecular modeling suggests a “sandwich” mode of binding and the flexibility of the dinculear motif can allow design for more selective and stronger-binding complexes. Based on these results a further prototype, [{Pt(en)(9-EtGua)}2μ-H2N(CH2)6NH2]4+, incorporating the purine 9-Ethylguanine (9-EtG) as a stacking moiety was prepared which showed good cytotoxicity in A2780 and OsACL tumor cell lines.
Zinc ions and copper ions are known to play important and contrasting roles in apoptosis, with zinc inhibiting the pro‐apoptotic protein caspase‐3. In their Communication on , N. P. Farrell and co‐workers present evidence that zinc inhibits caspase‐3 through interacting with the active‐site histidine residue rather than with the catalytic dyad as had previously been assumed.
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