Goran Kragol - Academia.edu (original) (raw)
Papers by Goran Kragol
Journal of Medicinal Chemistry, 2011
Macrolides with 14- and 15-membered ring are characterized by high and extensive tissue distribut... more Macrolides with 14- and 15-membered ring are characterized by high and extensive tissue distribution, as well as good cellular accumulation and retention. Since macrolide structures do not fit the Lipinski rule of five, macrolide pharmacokinetic properties cannot be successfully predicted by common models based on data for small molecules. Here we describe the development of the first models for macrolide cellular pharmacokinetics. By comparison of cellular accumulation and retention in six human primary cell cultures of leukocytic and lung origin, as well as in lung carcinoma cell line NCI-H292, this cell line was found to be an adequate representative cell type for modeling macrolide cellular pharmacokinetics. Accumulation and retention in the NCI-H292 cells, as well as various physicochemical properties, were determined for a set of 48 rationally designed basic macrolide compounds. Classification models for predicting macrolide cellular accumulation and retention were developed using relatively easily determined and conceptually simple descriptors: experimentally determined physicochemical parameters ChromlogD and CHI IAM, as well as a calculated number of positively charged atoms (POS). The models were further tested and improved by addition of 37 structurally diverse macrolide molecules.
European Journal of Medicinal Chemistry, 2011
18-crown-6 ethers are known to exert their biological activity by transporting K + ions across ce... more 18-crown-6 ethers are known to exert their biological activity by transporting K + ions across cell membranes. Using non-linear Support Vector Machines regression, we searched for structural features that influence antiproliferative activity in a diverse set of 19 known oxa-, monoaza-and diaza-18-crown-6 ethers. Here, we show that the logP of the molecule is the most important molecular descriptor, among ~1300 tested descriptors, in determining biological potency (R 2 cv =0.704). The optimal logP was at 5.5 (Ghose-Crippen ALOGP estimate) while both higher and lower values were detrimental to biological potency. After controlling for logP, we found that the antiproliferative activity of the molecule was generally not affected by side chain length, molecular symmetry, or presence of side chain amide links.
Pharmacological Research, 2012
Exceptional therapeutic effects of macrolides in treating various infections and inflammatory con... more Exceptional therapeutic effects of macrolides in treating various infections and inflammatory conditions can be significantly contributed to their unique pharmacokinetic properties. Macrolides accumulate in cells and tissues, with concentrations usually 10 to more than 100 times higher of those measured in plasma. Intracellular distribution of macrolides has so far been examined using extensive subcellular fractionation techniques, radiolabeled compounds and conventional pharmacokinetic methods. In this study we evaluated four fluorescently labeled macrolides on their applicability to monitor azithromycin distribution in vitro and in vivo. 9-Deoxo-9a-{3-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]propyl}-9aaza-9a-homoerythromycin A (9a-NBD-azithromycin) was selected as a compound with most similar cellular pharmacokinetics to azithromycin. 9a-NBD-azithromycin demonstrated antimicrobial properties comparable to azithromycin, displayed the same biological activity profile in LPS-stimulated J774A.1 murine macrophage cells and, even though it accumulated in cells almost 50% more than azithromycin, it showed same rate of retention. Identical to azithromycin, 9a-NBD-azithromycin was localized in lysosomes of J774A.1 cells.
Bioorganic & Medicinal Chemistry, 2010
In this paper synthesis of macrolones 1-18 starting from azithromycin is reported. Two key steps ... more In this paper synthesis of macrolones 1-18 starting from azithromycin is reported. Two key steps in the construction of the linker between macrolide and quinolone moiety, are formation of central ether bond by alkylation of unactivated OH group, and formation of terminal C-C bond at 6-position of the quinolone unit. Due to the difficulty in formation of these two bonds the study of alternative synthetic methodologies and optimization of the conditions for the selected routes was required. Formation of C-4 00 -O-ether bond was completed by modified Michael addition, whereas O-alkylation via diazonium cation proved to be the most effective in formation of the central allylic or propargylic ether bond. Comparison of Heck and Sonogashira reaction revealed the former as preferred route to the C-C bond formation at C(6) position of the quinolone unit. Most of the target compounds exhibited highly favorable antibacterial activity against common respiratory pathogens, without significant cytotoxicity profile when tested in vitro on eukaryotic cell lines.
Pharmacological Research, 2012
Exceptional therapeutic effects of macrolides in treating various infections and inflammatory con... more Exceptional therapeutic effects of macrolides in treating various infections and inflammatory conditions can be significantly contributed to their unique pharmacokinetic properties. Macrolides accumulate in cells and tissues, with concentrations usually 10 to more than 100 times higher of those measured in plasma. Intracellular distribution of macrolides has so far been examined using extensive subcellular fractionation techniques, radiolabeled compounds and conventional pharmacokinetic methods. In this study we evaluated four fluorescently labeled macrolides on their applicability to monitor azithromycin distribution in vitro and in vivo. 9-Deoxo-9a-{3-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]propyl}-9aaza-9a-homoerythromycin A (9a-NBD-azithromycin) was selected as a compound with most similar cellular pharmacokinetics to azithromycin. 9a-NBD-azithromycin demonstrated antimicrobial properties comparable to azithromycin, displayed the same biological activity profile in LPS-stimulated J774A.1 murine macrophage cells and, even though it accumulated in cells almost 50% more than azithromycin, it showed same rate of retention. Identical to azithromycin, 9a-NBD-azithromycin was localized in lysosomes of J774A.1 cells.
Journal of Molecular Structure, 2000
The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K... more The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K]+(C6H2N3O7)−;} have been studied by means of NMR-spectroscopy, X-ray structure analysis and molecular mechanics calculations. In the gas phase and in the CDCl3 solution, in both 1 and [1-K]+ the most stable conformation of C2h symmetry is found. However, in the solid state 1 adopts crystallographic Ci symmetry while its potassium picrato complex exhibits C1 symmetry.
Zeitschrift für Kristallographie - New Crystal Structures, 2000
The training data includes 25 chemical compounds with measured SW620 (colon), MCF-7 (breast) and ... more The training data includes 25 chemical compounds with measured SW620 (colon), MCF-7 (breast) and H460 (lung). In addition to the antitumor effects, we measured bacteriostatic All compounds are crown ethers, derivatives of 18-crown-6 (including oxa and aza-crowns), frequently carrying adamantane or other hydrophobic substituents. 18-crown-6 are known to shuttle potassium ions across biological membranes.
Tetrahedron, 1997
... Cs+ picrates that are strikingly similar to those shown by 15crown5.17 However, an important ... more ... Cs+ picrates that are strikingly similar to those shown by 15crown5.17 However, an important distinction is that 10 displayed 33 greater avidity toward Nat vish vis 15crown5 ... Mehta, G.; Rao, KS; Krishnamurthy, N.; Srinivas, V.; Balasubramanian, D. Tetrahedron 1989, 45, 2743. ...
European Journal of Organic Chemistry, 1999
ABSTRACT
Journal of Molecular Structure, 2000
The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K... more The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K]+(C6H2N3O7)−;} have been studied by means of NMR-spectroscopy, X-ray structure analysis and molecular mechanics calculations. In the gas phase and in the CDCl3 solution, in both 1 and [1-K]+ the most stable conformation of C2h symmetry is found. However, in the solid state 1 adopts crystallographic Ci symmetry while its potassium picrato complex exhibits C1 symmetry.
European Journal of Organic Chemistry, 2003
ABSTRACT
Bioorganic & Medicinal Chemistry, 2012
A set of 8-methylene-, 8-methyl-, and 8-methyl-9-dihydro-oleandomycin derivatives having differen... more A set of 8-methylene-, 8-methyl-, and 8-methyl-9-dihydro-oleandomycin derivatives having different combinations of stereochemistries at positions C-8 and/or C-9 have been prepared in a chemoselective and stereoselective manner and tested in vitro for antibacterial activity and inhibition of IL-6 production. Configurations of the stereocenters at C-8 and C-9 were determined using 2D NMR techniques. We have shown that change of stereochemistry at these positions can exert a major influence on antibacterial activity as well as IL-6 inhibition, providing novel macrolide derivatives with diminished antibacterial and potent anti-inflammatory activity. In addition, the anti-inflammatory activity observed in vitro was confirmed in an in vivo model of lipopolysaccharide-induced inflammation.
Tetrahedron, 2001
AbstractÐThe synthesis of a series of adamantane-and 2-oxaadamantane-functionalized crown ethers ... more AbstractÐThe synthesis of a series of adamantane-and 2-oxaadamantane-functionalized crown ethers 1±7 is described. Alkali metal picrate extraction pro®les have been determined for these novel ionophores. The ability of crown ethers 1±7 to extract the alkali metal picrates was compared with that of benzo-15-crown-5 and 18-crown-6. Also, Na 1 -and K 1 -transport, the ability of ionophores l±3 to transport Na 1 -and K 1 across a bulk liquid membrane was measured. The results of alkali metal cation extraction experiments showed that the complexation properties of crown ethers 1 and 2 are comparable to that of benzo-15-crown-5 and 18-crown-6, respectively. Crown ether 3 showed enhanced extractability for all cations but lower selectivity compared to 18-crown-6. However, adamantano-crown ethers 4±7 showed almost negligible extraction capability toward any of the alkali cations. The observed differences among the complexation abilities of the ionophores 1±3 and 4±7 are rationalized on the basis of the results of a molecular modeling study of their corresponding K 1 complexes. q
Tetrahedron, 2001
The development of peptide-based drugs requires the chemical synthesis of systems as complex as t... more The development of peptide-based drugs requires the chemical synthesis of systems as complex as they appear in nature. Most bioactive peptides have to be associated with co-activators and delivery or targeting domains, the synthesis of such complexes is far from trivial. In efforts to develop a prototype for a new generation of peptide vaccines, a peptide construct was prepared, using
Tetrahedron, 1998
... In addition, we thank Professor Jennifer S. Brodbelt (Department of Chemistry, University of ... more ... In addition, we thank Professor Jennifer S. Brodbelt (Department of Chemistry, University of Texas at" Austin) for having kindly obtained the high-resolution chemical ionization mass spectral data reported herein.9696 AP Marchand et al. ... GR Newcome and JM Robinson, J. Chem ...
Angewandte Chemie International Edition, 2005
ABSTRACT
Biochemistry, 2001
Recently, we documented that the short, proline-rich antibacterial peptides pyrrhocoricin, drosoc... more Recently, we documented that the short, proline-rich antibacterial peptides pyrrhocoricin, drosocin, and apidaecin interact with the bacterial heat shock protein DnaK, and peptide binding to DnaK can be correlated with antimicrobial activity. In the current report we studied the mechanism of action of these peptides and their binding sites to Escherichia coli DnaK. Biologically active pyrrhocoricin made of L-amino acids diminished the ATPase activity of recombinant DnaK. The inactive D-pyrrhocoricin analogue and the membrane-active antibacterial peptide cecropin A or magainin 2 failed to inhibit the DnaK-mediated phosphate release from adenosine 5'-triphosphate (ATP). The effect of pyrrhocoricin on DnaK's other significant biological function, the refolding of misfolded proteins, was studied by assaying the alkaline phosphatase and beta-galactosidase activity of live bacteria. Remarkably, both enzyme activities were reduced upon incubation with L-pyrrhocoricin or drosocin. D-Pyrrhocoricin, magainin 2, or buforin II, an antimicrobial peptide involved in binding to bacterial nucleic acids, had only negligible effect. According to fluorescence polarization and dot blot analysis of synthetic DnaK fragments and labeled pyrrhocoricin analogues, pyrrhocoricin bound with a K(d) of 50.8 microM to the hinge region around the C-terminal helices D and E, at the vicinity of amino acids 583 and 615. Pyrrhocoricin binding was not observed to the homologous DnaK fragment of Staphylococcus aureus, a pyrrhocoricin nonresponsive strain. In line with the lack of ATPase inhibition, drosocin binding appears to be slightly shifted toward the D helix. Our data suggest that drosocin and pyrrhocoricin binding prevents the frequent opening and closing of the multihelical lid over the peptide-binding pocket of DnaK, permanently closes the cavity, and inhibits chaperone-assisted protein folding. The biochemical results were strongly supported by molecular modeling of DnaK-pyrrhocoricin interactions. Due to the prominent sequence variations of procaryotic and eucaryotic DnaK molecules in the multihelical lid region, our findings pave the road for the design of strain-specific antibacterial peptides and peptidomimetics. Far-fetched applications of the species-specific inhibition of chaperone-assisted protein folding include the control of not only bacteria but also fungi, parasites, insects, and perhaps rodents.
Journal of Medicinal Chemistry, 2006
A series of new peptides (8-25) containing different unnatural amino acids of the adamantane type... more A series of new peptides (8-25) containing different unnatural amino acids of the adamantane type (1-6), was synthesized. Possible cytotoxic activity on human cervical adenocarcinoma (HeLa), larynx carcinoma (HEp-2), colon carcinomas (HT-29, Caco-2), poorly differentiated cells from lymph node metastasis of colon carcinoma (SW-620), mammary gland adenocarcinoma (MCF-7), and melanoma (HBL) cells were tested by the MTT assay. The results were compared with the effect of methionine-enkephalin (Tyr-Gly-Gly-Phe-Met, or opioid growth factor, OGF), and its shorter N-terminal fragments. Peptide analogues containing C(alpha alpha)-dialkylated glycine (Aaa1, 1) or C(alpha)-alkylated glycine (Aaa2, 2) amino acid residues showed antitumor activity against melanoma, larynx carcinoma, colon carcinomas, and colon metastasis cell lines in vitro. The pentapeptide Tyr-(R,S)-Aaa2-Gly-Phe-Met (18) was the most effective analogue especially against the most antitumor drug-resistant cell lines HEp-2 and SW-620. Apoptosis as a mode of cell death was confirmed in these tumor cells after exposure to pentapeptide 18.
Journal of Medicinal Chemistry, 2011
Macrolides with 14- and 15-membered ring are characterized by high and extensive tissue distribut... more Macrolides with 14- and 15-membered ring are characterized by high and extensive tissue distribution, as well as good cellular accumulation and retention. Since macrolide structures do not fit the Lipinski rule of five, macrolide pharmacokinetic properties cannot be successfully predicted by common models based on data for small molecules. Here we describe the development of the first models for macrolide cellular pharmacokinetics. By comparison of cellular accumulation and retention in six human primary cell cultures of leukocytic and lung origin, as well as in lung carcinoma cell line NCI-H292, this cell line was found to be an adequate representative cell type for modeling macrolide cellular pharmacokinetics. Accumulation and retention in the NCI-H292 cells, as well as various physicochemical properties, were determined for a set of 48 rationally designed basic macrolide compounds. Classification models for predicting macrolide cellular accumulation and retention were developed using relatively easily determined and conceptually simple descriptors: experimentally determined physicochemical parameters ChromlogD and CHI IAM, as well as a calculated number of positively charged atoms (POS). The models were further tested and improved by addition of 37 structurally diverse macrolide molecules.
European Journal of Medicinal Chemistry, 2011
18-crown-6 ethers are known to exert their biological activity by transporting K + ions across ce... more 18-crown-6 ethers are known to exert their biological activity by transporting K + ions across cell membranes. Using non-linear Support Vector Machines regression, we searched for structural features that influence antiproliferative activity in a diverse set of 19 known oxa-, monoaza-and diaza-18-crown-6 ethers. Here, we show that the logP of the molecule is the most important molecular descriptor, among ~1300 tested descriptors, in determining biological potency (R 2 cv =0.704). The optimal logP was at 5.5 (Ghose-Crippen ALOGP estimate) while both higher and lower values were detrimental to biological potency. After controlling for logP, we found that the antiproliferative activity of the molecule was generally not affected by side chain length, molecular symmetry, or presence of side chain amide links.
Pharmacological Research, 2012
Exceptional therapeutic effects of macrolides in treating various infections and inflammatory con... more Exceptional therapeutic effects of macrolides in treating various infections and inflammatory conditions can be significantly contributed to their unique pharmacokinetic properties. Macrolides accumulate in cells and tissues, with concentrations usually 10 to more than 100 times higher of those measured in plasma. Intracellular distribution of macrolides has so far been examined using extensive subcellular fractionation techniques, radiolabeled compounds and conventional pharmacokinetic methods. In this study we evaluated four fluorescently labeled macrolides on their applicability to monitor azithromycin distribution in vitro and in vivo. 9-Deoxo-9a-{3-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]propyl}-9aaza-9a-homoerythromycin A (9a-NBD-azithromycin) was selected as a compound with most similar cellular pharmacokinetics to azithromycin. 9a-NBD-azithromycin demonstrated antimicrobial properties comparable to azithromycin, displayed the same biological activity profile in LPS-stimulated J774A.1 murine macrophage cells and, even though it accumulated in cells almost 50% more than azithromycin, it showed same rate of retention. Identical to azithromycin, 9a-NBD-azithromycin was localized in lysosomes of J774A.1 cells.
Bioorganic & Medicinal Chemistry, 2010
In this paper synthesis of macrolones 1-18 starting from azithromycin is reported. Two key steps ... more In this paper synthesis of macrolones 1-18 starting from azithromycin is reported. Two key steps in the construction of the linker between macrolide and quinolone moiety, are formation of central ether bond by alkylation of unactivated OH group, and formation of terminal C-C bond at 6-position of the quinolone unit. Due to the difficulty in formation of these two bonds the study of alternative synthetic methodologies and optimization of the conditions for the selected routes was required. Formation of C-4 00 -O-ether bond was completed by modified Michael addition, whereas O-alkylation via diazonium cation proved to be the most effective in formation of the central allylic or propargylic ether bond. Comparison of Heck and Sonogashira reaction revealed the former as preferred route to the C-C bond formation at C(6) position of the quinolone unit. Most of the target compounds exhibited highly favorable antibacterial activity against common respiratory pathogens, without significant cytotoxicity profile when tested in vitro on eukaryotic cell lines.
Pharmacological Research, 2012
Exceptional therapeutic effects of macrolides in treating various infections and inflammatory con... more Exceptional therapeutic effects of macrolides in treating various infections and inflammatory conditions can be significantly contributed to their unique pharmacokinetic properties. Macrolides accumulate in cells and tissues, with concentrations usually 10 to more than 100 times higher of those measured in plasma. Intracellular distribution of macrolides has so far been examined using extensive subcellular fractionation techniques, radiolabeled compounds and conventional pharmacokinetic methods. In this study we evaluated four fluorescently labeled macrolides on their applicability to monitor azithromycin distribution in vitro and in vivo. 9-Deoxo-9a-{3-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]propyl}-9aaza-9a-homoerythromycin A (9a-NBD-azithromycin) was selected as a compound with most similar cellular pharmacokinetics to azithromycin. 9a-NBD-azithromycin demonstrated antimicrobial properties comparable to azithromycin, displayed the same biological activity profile in LPS-stimulated J774A.1 murine macrophage cells and, even though it accumulated in cells almost 50% more than azithromycin, it showed same rate of retention. Identical to azithromycin, 9a-NBD-azithromycin was localized in lysosomes of J774A.1 cells.
Journal of Molecular Structure, 2000
The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K... more The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K]+(C6H2N3O7)−;} have been studied by means of NMR-spectroscopy, X-ray structure analysis and molecular mechanics calculations. In the gas phase and in the CDCl3 solution, in both 1 and [1-K]+ the most stable conformation of C2h symmetry is found. However, in the solid state 1 adopts crystallographic Ci symmetry while its potassium picrato complex exhibits C1 symmetry.
Zeitschrift für Kristallographie - New Crystal Structures, 2000
The training data includes 25 chemical compounds with measured SW620 (colon), MCF-7 (breast) and ... more The training data includes 25 chemical compounds with measured SW620 (colon), MCF-7 (breast) and H460 (lung). In addition to the antitumor effects, we measured bacteriostatic All compounds are crown ethers, derivatives of 18-crown-6 (including oxa and aza-crowns), frequently carrying adamantane or other hydrophobic substituents. 18-crown-6 are known to shuttle potassium ions across biological membranes.
Tetrahedron, 1997
... Cs+ picrates that are strikingly similar to those shown by 15crown5.17 However, an important ... more ... Cs+ picrates that are strikingly similar to those shown by 15crown5.17 However, an important distinction is that 10 displayed 33 greater avidity toward Nat vish vis 15crown5 ... Mehta, G.; Rao, KS; Krishnamurthy, N.; Srinivas, V.; Balasubramanian, D. Tetrahedron 1989, 45, 2743. ...
European Journal of Organic Chemistry, 1999
ABSTRACT
Journal of Molecular Structure, 2000
The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K... more The structure of bis[(1,3)2-oxaadamantano]-18-crown-6 (1) and its potassium picrato complex {[1-K]+(C6H2N3O7)−;} have been studied by means of NMR-spectroscopy, X-ray structure analysis and molecular mechanics calculations. In the gas phase and in the CDCl3 solution, in both 1 and [1-K]+ the most stable conformation of C2h symmetry is found. However, in the solid state 1 adopts crystallographic Ci symmetry while its potassium picrato complex exhibits C1 symmetry.
European Journal of Organic Chemistry, 2003
ABSTRACT
Bioorganic & Medicinal Chemistry, 2012
A set of 8-methylene-, 8-methyl-, and 8-methyl-9-dihydro-oleandomycin derivatives having differen... more A set of 8-methylene-, 8-methyl-, and 8-methyl-9-dihydro-oleandomycin derivatives having different combinations of stereochemistries at positions C-8 and/or C-9 have been prepared in a chemoselective and stereoselective manner and tested in vitro for antibacterial activity and inhibition of IL-6 production. Configurations of the stereocenters at C-8 and C-9 were determined using 2D NMR techniques. We have shown that change of stereochemistry at these positions can exert a major influence on antibacterial activity as well as IL-6 inhibition, providing novel macrolide derivatives with diminished antibacterial and potent anti-inflammatory activity. In addition, the anti-inflammatory activity observed in vitro was confirmed in an in vivo model of lipopolysaccharide-induced inflammation.
Tetrahedron, 2001
AbstractÐThe synthesis of a series of adamantane-and 2-oxaadamantane-functionalized crown ethers ... more AbstractÐThe synthesis of a series of adamantane-and 2-oxaadamantane-functionalized crown ethers 1±7 is described. Alkali metal picrate extraction pro®les have been determined for these novel ionophores. The ability of crown ethers 1±7 to extract the alkali metal picrates was compared with that of benzo-15-crown-5 and 18-crown-6. Also, Na 1 -and K 1 -transport, the ability of ionophores l±3 to transport Na 1 -and K 1 across a bulk liquid membrane was measured. The results of alkali metal cation extraction experiments showed that the complexation properties of crown ethers 1 and 2 are comparable to that of benzo-15-crown-5 and 18-crown-6, respectively. Crown ether 3 showed enhanced extractability for all cations but lower selectivity compared to 18-crown-6. However, adamantano-crown ethers 4±7 showed almost negligible extraction capability toward any of the alkali cations. The observed differences among the complexation abilities of the ionophores 1±3 and 4±7 are rationalized on the basis of the results of a molecular modeling study of their corresponding K 1 complexes. q
Tetrahedron, 2001
The development of peptide-based drugs requires the chemical synthesis of systems as complex as t... more The development of peptide-based drugs requires the chemical synthesis of systems as complex as they appear in nature. Most bioactive peptides have to be associated with co-activators and delivery or targeting domains, the synthesis of such complexes is far from trivial. In efforts to develop a prototype for a new generation of peptide vaccines, a peptide construct was prepared, using
Tetrahedron, 1998
... In addition, we thank Professor Jennifer S. Brodbelt (Department of Chemistry, University of ... more ... In addition, we thank Professor Jennifer S. Brodbelt (Department of Chemistry, University of Texas at" Austin) for having kindly obtained the high-resolution chemical ionization mass spectral data reported herein.9696 AP Marchand et al. ... GR Newcome and JM Robinson, J. Chem ...
Angewandte Chemie International Edition, 2005
ABSTRACT
Biochemistry, 2001
Recently, we documented that the short, proline-rich antibacterial peptides pyrrhocoricin, drosoc... more Recently, we documented that the short, proline-rich antibacterial peptides pyrrhocoricin, drosocin, and apidaecin interact with the bacterial heat shock protein DnaK, and peptide binding to DnaK can be correlated with antimicrobial activity. In the current report we studied the mechanism of action of these peptides and their binding sites to Escherichia coli DnaK. Biologically active pyrrhocoricin made of L-amino acids diminished the ATPase activity of recombinant DnaK. The inactive D-pyrrhocoricin analogue and the membrane-active antibacterial peptide cecropin A or magainin 2 failed to inhibit the DnaK-mediated phosphate release from adenosine 5'-triphosphate (ATP). The effect of pyrrhocoricin on DnaK's other significant biological function, the refolding of misfolded proteins, was studied by assaying the alkaline phosphatase and beta-galactosidase activity of live bacteria. Remarkably, both enzyme activities were reduced upon incubation with L-pyrrhocoricin or drosocin. D-Pyrrhocoricin, magainin 2, or buforin II, an antimicrobial peptide involved in binding to bacterial nucleic acids, had only negligible effect. According to fluorescence polarization and dot blot analysis of synthetic DnaK fragments and labeled pyrrhocoricin analogues, pyrrhocoricin bound with a K(d) of 50.8 microM to the hinge region around the C-terminal helices D and E, at the vicinity of amino acids 583 and 615. Pyrrhocoricin binding was not observed to the homologous DnaK fragment of Staphylococcus aureus, a pyrrhocoricin nonresponsive strain. In line with the lack of ATPase inhibition, drosocin binding appears to be slightly shifted toward the D helix. Our data suggest that drosocin and pyrrhocoricin binding prevents the frequent opening and closing of the multihelical lid over the peptide-binding pocket of DnaK, permanently closes the cavity, and inhibits chaperone-assisted protein folding. The biochemical results were strongly supported by molecular modeling of DnaK-pyrrhocoricin interactions. Due to the prominent sequence variations of procaryotic and eucaryotic DnaK molecules in the multihelical lid region, our findings pave the road for the design of strain-specific antibacterial peptides and peptidomimetics. Far-fetched applications of the species-specific inhibition of chaperone-assisted protein folding include the control of not only bacteria but also fungi, parasites, insects, and perhaps rodents.
Journal of Medicinal Chemistry, 2006
A series of new peptides (8-25) containing different unnatural amino acids of the adamantane type... more A series of new peptides (8-25) containing different unnatural amino acids of the adamantane type (1-6), was synthesized. Possible cytotoxic activity on human cervical adenocarcinoma (HeLa), larynx carcinoma (HEp-2), colon carcinomas (HT-29, Caco-2), poorly differentiated cells from lymph node metastasis of colon carcinoma (SW-620), mammary gland adenocarcinoma (MCF-7), and melanoma (HBL) cells were tested by the MTT assay. The results were compared with the effect of methionine-enkephalin (Tyr-Gly-Gly-Phe-Met, or opioid growth factor, OGF), and its shorter N-terminal fragments. Peptide analogues containing C(alpha alpha)-dialkylated glycine (Aaa1, 1) or C(alpha)-alkylated glycine (Aaa2, 2) amino acid residues showed antitumor activity against melanoma, larynx carcinoma, colon carcinomas, and colon metastasis cell lines in vitro. The pentapeptide Tyr-(R,S)-Aaa2-Gly-Phe-Met (18) was the most effective analogue especially against the most antitumor drug-resistant cell lines HEp-2 and SW-620. Apoptosis as a mode of cell death was confirmed in these tumor cells after exposure to pentapeptide 18.