Aldona Karaczyn

Studies on the interactions between human serum albumin and trans-indazolium (bisindazole) tetrachlororuthenate(III)

The interactions between HInd[RuInd2Cl4] and human serum albumin have been investigated through UV-Vis, circular dichroism (CD), fluorescence spectroscopy and the inductively coupled plasma-atomic emission spectroscopy (ICP(AES)) method. Binding of Ru(III)-indazole species to albumin has strong impact on protein structure and it influences considerably albumin binding of other molecules like warfarin, heme or metal ions. The metal complex-human serum albumin (HAS) interactions cause conformational changes with loss of helical stability of the protein and local perturbation in the domain IIA binding pocket. The relative fluorescence intensity of the ruthenium-bound HSA decreased, suggesting that perturbation around the Trp 214 residue took place. This was confirmed by the destabilization of the warfarin-binding site, which includes Trp 214, observed in the metal-bound HSA.

Regulation of hypoxia-inducible genes by ETS1 transcription factor

Hypoxia-inducible factor (HIF-1) regulates the expression of genes that facilitate tumor cell survival by making them more resistant to therapeutic intervention. Recent evidence suggests that the activation of other transcription factors, in cooperation with HIF-1 or acting alone, is involved in the upregulation of hypoxia-inducible genes. Here we report that high cell density, a condition that might mimic the physiologic situation in growing tumor and most probably representing nutritional starvation, upregulates hypoxia-inducible genes. This upregulation can occur in HIF-independent manner since hypoxia-inducible genes carbonic anhydrase 9 (CA9), lysyloxidase like 2 (LOXL2) and n-myc-down regulated 1 (NDRG1)/calcium activated protein (Cap43) can be upregulated by increased cell density under both normoxic and hypoxic conditions in both HIF-1 alpha-proficient and -deficient mouse fibroblasts. Moreover, cell density upregulates the same genes in 1HAEo- and A549 human lung epithelial cells. Searching for other transcription factors involved in the regulation of hypoxia-inducible genes by cell density, we focused our attention on ETS1. As reported previously, members of v-ets erythroblastosis virus E26 oncogene homolog (ETS) family transcription factors participate in the upregulation of hypoxia-inducible genes. Here, we provide evidence that ETS1 protein is upregulated at high cell density in both human and mouse cells. The involvement of ETS1 in the upregulation of hypoxia-inducible genes was further confirmed in a luciferase reporter assay using cotransfection of ETS1 expression vector with NDRG1/Cap43 promoter construct. The downregulation of ETS1 expression with small interfering RNA (siRNA) inhibited the upregulation of CA9 and NDRG1/Cap43 caused by increased cell density. Collectively, our data indicate the involvement of ETS1 along with HIF-1 in regulating hypoxia-inducible genes.

COPPER(II) BINDING TO TOBRAMYCIN : POTENTIOMETRIC AND SPECTROSCOPIC STUDIES

Protonation and Cu(II) binding by tobramycin, an aminoglycosidic antibiotic, was studied by potentiometry and UV-vis, CD and EPR spectroscopies. A range of mononuclear complexes of a general formula CuHnL was found, with n between 3 and -2. Tobramycin anchors Cu(II) with an ¿NH2, O-¿ chelate of the C-ring of its molecule. The amino and hydroxyl groups of the A-ring of tobramycin also participate in the binding at pH 7 and higher. The resulting structure involves both terminal aminosugar rings but eliminates the donors of the central streptamine unit from the coordination. A comparison between tobramycin and its close analog, kanamycin B [M. Jezowska-Bojczuk, W. Bal and H. Kozłowski, Inorg. Chim. Acta, 275-276 (1998) 541-545] reveals the importance of the A3-OH group for the binding properties of these aminoglycosides.

Copper(II) binding to geneticin, a gentamycin analog

Abstract Protonation and Cu(II) binding by geneticin, an aminoglycosidic antibiotic of the gentamycin family, was studied by potentiometry and spectroscopic techniques (UV–vis, CD and EPR). Mononuclear complexes of a general formula CuH n L were found, with n between 2 and −2. In contrast to previously studied aminoglycosides, an additional complex with two aminoglycoside molecules, CuH −3 L 2 , was found above pH 9. Geneticin binds Cu(II) primarily with an {NHCH 3 , O − } chelate of the C-ring of its molecule. The amino and hydroxyl groups of the A-ring of geneticin also participate in the binding above pH 7. The presence of the methyl substituent on the C-ring amine nitrogen decreases Cu(II) affinity to this ring slightly compared to other aminoglycosides sharing homologous Cu(II) binding sites (tobramycin, kanamycin B). This is more than compensated by the increased affinity of the A-ring, due to the absence of a C-6 amine in geneticin.

Copper(II) and nickel(II) complexes with oxime analogues of amino acids. Potentiometric, spectroscopic and X-ray studies of complexes with 2-cyano-2-(hydroxyimino)acetic acid and its ethane-1,2-diamine derivative

2-Cyano-2-(hydroxyimino)acetic acid was found to be an effective ligand for Cu2+ and Ni2+ ions over a very broad pH range. Potentiometric and spectroscopic data indicate the formation of dimeric complexes with Cu2+ ions and the first X-ray evidence for dimer formation is obtained. Conjugation of the oximic moiety with ethane-1,2-diamine leads to another very effective family of chelating agents, although metal ion co-ordination may induce amide bond hydrolysis as indicated by the crystal structure.

Co-ordination of copper(II) and nickel(II) ions by a novel open chain oxime ligand

Potentiometric spectroscopic and X-ray studies of the open chain oxime ligand, N,N′-bis(2-hydroxyimino-propionyl)propane-1,3-diamine and its complexes with NiII and CuII ions showed a very high efficacy of the ligand studied in the co-ordination of both CuII and NiII ions. The metal complexes are very stable and square-planar with four nitrogens involved in metal-ion binding. These complexes may be additionally stabilised by hydrogen bonds between two oxime oxygen atoms.

Co-ordination ability of novel tetradentate amide-and-oxime ligands: differential binding to CuII and NiII

Four cationic and anionic complexes of NiII and CuII with new open chain oxime-and-amide ligands, N,N′-bis(2-hydroxyiminopropionyl)-1,2-diaminoethane (H2pen) and 1,4-diaminobutane (H2pab) were isolated and characterised by X-ray crystallography and a variety of spectroscopic techniques. The ligands were found to exhibit two types of co-ordination modes depending on pH: N(oxime),O(amide) bis-bidentate and N2(oxime),N2(amide) tetradentate. The first bonding mode leads to bridging of the two ligands thus forming binuclear cationic complexes [{Cu(Hpab)(ClO4)·2H2O}2] 1b and Li[Ni(H–1pen)]·4H2O 2a which have been determined by single-crystal X-ray analysis. In 1b the bis-bidentate co-ordination mode leads to the formation of a dimeric complex cation containing a loose 22-membered macrocyclic cavity incorporating two CuII ions as a part of the ring skeleton. The central atoms in 1b are distorted square-pyramidal with the water molecule in the apical position. N4-tetradentate co-ordination observed in 2a leads to the formation of a square-planar complex anion with a closed pseudo-macrocyclic environment stabilised by a short intramolecular H-bond between the oxime oxygen atoms.

A short intramolecular hydrogen bond is a key factor in the self-assembly of a dimeric complex with a 22-membered metallamacrocyclic cavity

The reaction of copper(II) perchlorate with the new open chain oxime–amide ligand, N,N′-bis(2-hydroxyiminopropionyl)butane-1,4-diamine (H2pab) in aqueous solution produced the dimeric complex [Cu(Hpab)]2[ClO4]2·4H2O containing a 22-membered macrocyclic cavity incorporating two copper(II) ions as a part of the ring framework.

CRYSTAL AND MOLECULAR STRUCTURE OF TWO TETRADENTATE OXIME-AND-AMIDE LIGANDS

Syntheses and X-ray structure analyses of two new tetradentate ligands with “oxime-and-amide” donor systems (N,N′-bis(2-hydroiminopropionyl)-1,2-diaminoethane and N,N′-bis(2- hydroxyiminopropionyl)-1,4-diaminobutane) were performed. The overall conformation of both ligands is distinctly different from that reported earlier for N,N′-bis(2-hydroxyiminopropionyl)- 1,2-diaminopropane. The number of methylene groups critically influences the ligand geometry and may have distinct impact on the co-ordinating ability of the ligands.

Insertion of oximic and hydroxamic functions into one simple amino acid creates a new family of powerful chelating agents

2-(Hydroxyimino)propanohydroxamic acid, a derivative of alanine in which the amino group has been transformed into oxime and the carboxylic group into a hydroxamic function, is a more powerful chelating agent for Cu2+ and Ni2+ ions than alanine itself, its oximic or hydroxamic analogues.