RESUMO
A series of PdII complexes with bis-(2-pyridylmethyl)glycine as a ligand of formula [PdX(bis-(2-pyridylmethyl)glycine)] where Xâ¯=â¯Cl, Br, I were prepared and the effect of the halogen nature in the antitumor activity of eight tumorigenic and one non-tumorigenic cell line was evaluated. The chloride derivative was further functionalized with a transferrin receptor binding peptide, generating the first PdII based metallopeptide. Its antitumor activity was also evaluated. However, among all the complexes, the chloride and iodine parent compounds showed the lowest GI50 values in the panel evaluated, and lowest GI50 than cisplatin in several cell lines. In contrast, the bromine derivative showed higher values of GI50 than chloride and iodine (around 30 - 50⯵M). The same trend was observed for the bovine serum albumin binding constant with higher values for iodine, chlorine, and bromine in this order. In aqueous solution, the chloride is exchanged by water while the bromine and iodine are not. DNA was evaluated as a target and showed no significative interaction for all the compounds. The results suggest sulfur-rich proteins and not DNA as a target. This report represents the first PdII metallopeptide reported, its evaluation in solution and antitumor activity. This work opens the possibilities for further functionalization of PdII complexes and the importance of the halogen coordination in the design of novel metallodrugs.
Assuntos
Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Halogênios/química , Paládio/química , Peptídeos/química , Peptídeos/farmacologia , Proteínas de Ligação a Transferrina/química , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Complexos de Coordenação/síntese química , Células HT29 , Humanos , Células MCF-7 , Espectroscopia de Ressonância Magnética , Peptídeos/síntese química , Relação Estrutura-Atividade , Difração de Raios XRESUMO
Entamoeba histolytica is an enteric protozoan that exclusively infects human beings. This parasite requires iron for its metabolic functions. Lactoferrin is a mammalian glycoprotein that chelates extracellular iron on mucosal surfaces, including the surface of the large intestine, where E. histolytica initiates infection. This work examined the interaction in vitro of E. histolytica trophozoites with human hololactoferrin (iron-saturated lactoferrin). A minimum concentration of 50 microM Fe from hololactoferrin supported growth of the amoeba. Amoebic binding sites for hololactoferrin were different from those for human apolactoferrin, holotransferrin and haemoglobin. One amoebic hololactoferrrin-binding polypeptide of 90 kDa was found, which was not observed after treatment of trophozoites with trypsin. Hololactoferrin-binding-protein levels increased in amoebas starved of iron, or grown in hololactoferrin. Internalization of hololactoferrin was inhibited by filipin. Endocytosed hololactoferrin colocalized with an anti-chick embryo caveolin mAb in amoebic vesicles, and lactoferrin was further detected in acidic vesicles; amoebic caveolin of 22 kDa was detected by Western blotting using this antibody. Cysteine proteases from amoebic extracts were able to cleave hololactoferrin. Together, these data indicate that E. histolytica trophozoites bind to hololactoferrin through specific membrane lactoferrin-binding proteins. This ferric protein might be internalized via caveolae-like microdomains, then used as an iron source, and degraded.
Assuntos
Anticorpos Antiprotozoários/imunologia , Endocitose/fisiologia , Entamoeba histolytica/metabolismo , Ferro/metabolismo , Lactoferrina/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Entamoeba histolytica/crescimento & desenvolvimento , Humanos , Proteínas de Ligação ao Ferro , Proteínas de Ligação a TransferrinaRESUMO
The transferrin-binding protein Bs (TbpBs) from the bacterium Neisseria meningitidis have been divided into two families according to genetic and antigenic features. TbpB from meningococcal strain B385 showed a molecular mass similar to that exhibited by TbpBs belonging to the high molecular mass family of TbpBs. TbpB was recognized by immunoassay using a specific serum directed against the TbpB of the reference strain for this family (strain M982). It was also recognized by a serum elicited against the TbpB of the reference strain for the low molecular mass family (strain B16B6). The tbpB gene from strain B385 was cloned and sequenced. The highest degree of sequence homology was found to be with the TbpBs belonging to the high molecular mass family, although a region of 14 amino acids that is only present in the TbpB from strain B16B6 was also found. This report illustrates a TbpB that shows hybrid antigenic and genetic behaviour.
Assuntos
Antígenos de Bactérias/genética , Proteínas de Transporte/genética , Neisseria meningitidis/genética , Sequência de Aminoácidos , Proteínas de Transporte/imunologia , Proteínas de Ligação ao Ferro , Dados de Sequência Molecular , Neisseria meningitidis/imunologia , Homologia de Sequência de Aminoácidos , Proteína B de Ligação a Transferrina , Proteínas de Ligação a TransferrinaRESUMO
The gene coding for the 98-kDa meningococcal outer membrane transferrin binding protein 1 (TbpA) from strain B385 was cloned and sequenced. Sequence comparison among its deduced aminoacid sequence and those from TbpA and the closely related LbpA (lactoferrin binding protein) gene from three different meningococcal strains, and four isolates from two other bacterial pathogens, showed that TbpA variability is confined to five specific segments, designated VR1 (199-287), VR2 (306-381), VR3 (480-546), VR4 (618-651) and VR5 (681-708). The third VR was the most variable among strains both at the nucleotide and amino acid levels. Six additional tbpA genes from different meningococcal strains were cloned and its VR3 sequence determined. On the basis of this data we were able to cluster tbpA genes in two groups: D (bearing a deletion in VR3) and N (nondeleted); all N and D strains belonging to the groups of high or low molecular weight transferrin receptor isotype, respectively. However, by phenogram analysis, the prototypical strain M982 (Group II) was clustered with M990 (B16B6 isotype, Group I). These results point to the existence of important exposed regions as well as to the possibility of horizontal gene exchange involving this locus. A topology model with 14 exposed loops and 28 membrane spanning segments was postulated. According to this tentative analysis, TbpA as well as LbpA proteins should form a gated channel in the neisserial outer membrane. The variable regions were located in the fifth, sixth, eighth, 10th and 11th loops respectively. Among TbpAs VR1, VR2, and VR3 resulted the most relevant regions.