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Markus Kaiser

Publications and source records attributed to Markus Kaiser.

12 recordsLinked to original sources
PubMed2020

Natural brominated phenoxyphenols kill persistent and biofilm-incorporated cells of MRSA and other pathogenic bacteria.

Due to a high unresponsiveness to chemotherapy, biofilm formation is an important medical problem that frequently occurs during infection with many bacterial pathogens. In this study, the marine sponge-derived natural compounds 4,6-dibromo-2-(2',4'-dibromophenoxy)phenol and 3,4,6-tribromo-2-(2',4'-dibromophenoxy)phenol were found to exhibit broad antibacterial activity against medically relevant gram-positive and gram-negative pathogens. The compounds were not only bactericidal against both replicating and stationary phase-persistent planktonic cells of methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa; they also killed biofilm-incorporated cells of both species while not affecting biofilm structural integrity. Moreover, these compounds were active against carbapenemase-producing Enterobacter sp. This simultaneous activity of compounds against different growth forms of both gram-positive and gram-negative bacteria is rare. Genome sequencing of spontaneous resistant mutants and proteome analysis suggest that resistance is mediated by downregulation of the bacterial EIIBC phosphotransferase components scrA and mtlA in MRSA likely leading to a lower uptake of the molecules. Due to their only moderate cytotoxicity against human cell lines, phenoxyphenols provide an interesting new scaffold for development of antimicrobial agents with activity against planktonic cells, persisters and biofilm-incoporated cells of ESKAPE pathogens. KEY POINTS: • Brominated phenoxyphenols kill actively replicating and biofilm-incorporated bacteria. • Phosphotransferase systems mediate uptake of brominated phenoxyphenols. • Downregulation of phosphotransferase systems mediate resistance.

Animals↗
PubMed2006

Neomycin-capped aromatic platforms: quadruplex DNA recognition and telomerase inhibition.

A series of aminoglycoside-capped macrocyclic structures has been prepared using intramolecular bis-tethering of neomycin on three aromatic platforms (phenanthroline, acridine, quinacridine). Based on NMR and calculations studies, it was found that the cyclic compounds adopt a highly flexible structure without conformational restriction of the aminoglycoside moiety. FRET-melting stabilization measurements showed that the series displays moderate to high affinity for the G4-conformation of human telomeric repeats, this effect being correlated with the size of the aromatic moiety. In addition, a FRET competition assay evidenced the poor binding ability of all macrocycles for duplex DNA and a clear binding preference for loop-containing intramolecular G4 structures compared to tetramolecular parallel G4 DNA. Finally, TRAP experiments demonstrated that the best G4-binder (quinacridine ) is also a potent and selective telomerase inhibitor with an IC(50) in the submicromolar range (200 nM).

Acridines↗
PubMed2006

Aminoglycoside-quinacridine conjugates: towards recognition of the P6.1 element of telomerase RNA.

A modular synthesis has been developed which allows easy and rapid attachment of one or two aminoglycoside units to a quinacridine intercalator, thereby leading to monomeric and dimeric conjugates. Melting temperature (Tm) experiments show that the tobramycin dimeric conjugate TD1 exhibits strong binding to the P6.1 element of human telomerase RNA. By contrast, tobramycin alone is much less efficient and the monomeric compound TM1 elicits a poor binding ability. Monitoring of the interaction by an electrophoretic mobility shift assay shows a 1:1 stoichiometry for the binding of the dimeric compound to the hairpin structure and confirms the lower affinity for a control duplex. Protection experiments with RNase T1 indicate interaction of the drug both in the stem and in the loop of the hairpin. Taken together, the data suggest a binding of TD1 inside the hairpin at the stem-loop junction. The same trends are observed with paromomycin and kanamycin analogues but with a lower affinity.

Acridines↗
PubMed2006

TMC-95-based inhibitor design provides evidence for the catalytic versatility of the proteasome.

TMC-95's natural cyclic tripeptide metabolites represent potent competitive proteasome inhibitors. The constrained conformation of TMC-95 proteasomal inhibitors provides the driving force for entropically high-affinity binding. Based on the crystal structure of the proteasome:TMC-95A complex, the synthetically challenging TMC-95 core structure was used for the design and synthesis of less demanding biphenyl-ether macrocycles, in which the biphenyl-ether moiety functions as an endocyclic clamp restricting its tripeptide backbone. These simplified analogs allowed us to identify high plasticity of the proteasomal tryptic-like specificity pocket. Biphenyl-ether compounds extended with an amide group were hydrolyzed by the proteasome, although the crystal structure of such proteasome:biphenyl-ether complexes revealed quenching of proteolysis at the acyl-enzyme intermediate. Our data reveal that biphenyl-ether derivatives bind noncovalently to the proteasomal tryptic-like active site in a reversible substrate-like manner without allosteric changes of active site residues.

Binding Sites↗
PubMed2004

Binding mode of TMC-95A analogues to eukaryotic 20S proteasome.

The complex thermodynamics that govern noncovalent protein-ligand interactions are still not fully understood, despite the exponential increase in experimental structural data available from X-ray crystallography and NMR spectroscopy. The eukaryotic 20S proteasome offers an ideal system for such studies as it contains in duplicate three proteolytically active sites with different substrate specificities. The natural product TMC-95A inhibits these proteolytic centers noncovalently with distinct affinities. X-ray crystallographic analysis of the complexes of the yeast proteasome core particle with this natural inhibitor and two synthetic analogues clearly revealed highly homologous hydrogen-bonding networks involving mainly the peptide backbone despite the strongly differentiated binding affinities to the three active sites of the 20S proteasome. The natural product and the two analogues are constrained in a rigid beta-type extended conformation by the endocyclic biaryl clamp, which preorganizes the peptide backbone for optimal adaptation of the ligands to the active site clefts and thus favors the binding processes entropically. However, the biaryl clamp also dictates the orientation of the P1 and P3 residues and their mode of interaction with the protein binding subsites. This limitation is optimally solved in TMC-95A with the conformationally restricted (Z)-prop-1-enyl group acting as P1 residue, at least for the chymotrypsin-like active site; however, it critically affects the inhibitory potencies of the analogues, thus suggesting the use of less-rigid endocyclic clamps in the design of proteasome inhibitors that allow for a better presentation of residues interacting with the active site clefts of the enzyme.

Binding Sites↗
PubMed2004

TMC-95A analogues with endocyclic biphenyl ether group as proteasome inhibitors.

TMC-95A, a cyclic tripeptide metabolite of Apiospora montagnei, is a potent competitive inhibitor of proteasome. Based on the X-ray structure of its complex with yeast proteasome, the synthetically challenging structure of this natural product was simplified in a first generation of analogues by replacing the highly oxidized side-chain biaryl system with a phenyl-oxindole group. In the present study, the TMC-95 biaryl group was substituted with a biphenyl ether with retainment of significant proteasome inhibition. Because of the facile synthetic access of tripeptides containing in i, i+2 positions residues of the isodityrosine type, this new generation of TMC-95 analogues may represent promising lead structures for further optimization of affinity and selectivity of proteasome inhibitors.

Biphenyl Compounds↗
PubMed2002

Navigation inside a protease: substrate selection and product exit in the tricorn protease from Thermoplasma acidophilum.

The proposed pathway and mechanism of substrate entry and product egress in the hexameric D3 symmetric tricorn protease from Thermoplasma acidophilum were explored by crystallographic studies of ligand complexes and by structure-based mutagenesis. Obstruction of the pore within the 7-bladed beta-propeller (beta7) domain by alkylation or oxidation of an engineered double cysteine mutant strongly decreased enzymatic activities. In line herewith, the crystal structure of the tricorn protease in complex with a trideca-peptide inhibitor modifying the catalytic Ser965 revealed part of the peptide trapped inside the channel of the beta7 domain. The cysteine mutation widening the lumen of the 6-bladed beta-propeller (beta6) domain enhanced catalytic activity, which was restored to normal values after its alkylation. A charge reversal mutant at the putative anchor site of the substrate C terminus, R131E-R132E, drastically reduced the proteolytic activity. The complex crystal structure of a peptide inhibitor with a diketo group at the cleavage site mapped the substrate recognition site and confirmed the role of Arg131-Arg132 as an anchor site. Our results strongly suggest the wider beta7 domain to serve as a selective filter and guide of the substrate to the sequestered active site, while the narrower beta6 domain routes the product to the surface. Moreover, we identified the role of Arg131-Arg132 in anchoring the substrate C terminus.

Alkylation↗
PubMed2002

Structural plasticity and noncovalent substrate binding in the GroEL apical domain. A study using electrospay ionization mass spectrometry and fluorescence binding studies.

Advances in understanding how GroEL binds to non-native proteins are reported. Conformational flexibility in the GroEL apical domain, which could account for the variety of substrates that GroEL binds, is illustrated by comparison of several independent crystallographic structures of apical domain constructs that show conformational plasticity in helices H and I. Additionally, ESI-MS indicates that apical domain constructs have co-populated conformations at neutral pH. To assess the ability of different apical domain conformers to bind co-chaperone and substrate, model peptides corresponding to the mobile loop of GroES and to helix D from rhodanese were studied. Analysis of apical domain-peptide complexes by ESI-MS indicates that only the folded or partially folded apical domain conformations form complexes that survive gas phase conditions. Fluorescence binding studies show that the apical domain can fully bind both peptides independently. No competition for binding was observed, suggesting the peptides have distinct apical domain-binding sites. Blocking the GroES-apical domain-binding site in GroEL rendered the chaperonin inactive in binding GroES and in assisting the folding of denatured rhodanese, but still capable of binding non-native proteins, supporting the conclusion that GroES and substrate proteins have, at least partially, distinct binding sites even in the intact GroEL tetradecamer.

Binding Sites↗
PubMed2002

Therapeutic effects of acetylsalicylic acid in giant cell arteritis.

OBJECTIVE: In giant cell arteritis (GCA), inflammatory lesions typically produce interferon-gamma(IFNgamma)-- and nuclear factor kappaB (NF-kappaB)-dependent monokines. Corticosteroids influence disease activity by repressing NF-kappaB-dependent genes but have only marginal effects on IFNgamma. The current study explored whether acetylsalicylic acid (ASA) had cytokine-repressing activity in GCA and could function as a steroid-sparing agent. METHODS: Temporal artery-severe combined immunodeficiency (SCID) mouse chimeras were created by engrafting inflamed temporal arteries into SCID mice. Chimeras were treated with ASA, indomethacin, or dexamethasone for 3 weeks. Temporal artery grafts were harvested and cytokine message was semiquantified by polymerase chain reaction-enzyme-linked immunosorbent assay. The ability of dexamethasone and ASA to suppress IFNgamma and interleukin-1beta (IL-1beta) messenger RNA and protein production was also tested in vitro using T cell clones and monocytes derived from patients with GCA. Drug-induced effects on the transcription factors NF-kappaB and activator protein 1 (AP-1) were assessed by electrophoretic mobility shift assays (EMSAs). RESULTS: At clinically relevant doses, 20-100 mg/kg, ASA was a highly effective inhibitor of cytokine transcription in temporal arteries. While dexamethasone preferentially targeted NF-kappaB-regulated monokines, ASA acted predominantly by suppressing IFNgamma. Indomethacin failed to reduce tissue IFNgamma transcription, which therefore excluded the inhibition of cyclooxygenases as a critical mechanism. IFNgamma production by T cell clones was highly sensitive to ASA-mediated suppression, whereas IL-1beta production by lipopolysaccharide-stimulated monocytes responded primarily to dexamethasone. The combination of ASA and dexamethasone had synergistic effects. EMSAs demonstrated that ASA interfered with the formation of AP-1, whereas dexamethasone suppressed the nuclear translocation of NF-kappaB. CONCLUSION: The results of this study provide evidence of the complementary action of ASA and corticosteroids in suppressing proinflammatory cytokines in the vascular lesions of GCA.

Animals↗