Urszula Uciechowska - Academia.edu (original) (raw)

Papers by Urszula Uciechowska

Research on DNA replication has not revealed the exact mechanism for replication complex de novo ... more Research on DNA replication has not revealed the exact mechanism for replication complex de novo assembly nor how the directionality of replication is determined. By using broad-host-range plasmid system we demonstrated evidence for direct involvement of a replication initiation protein in the process of polymerase recruitment. Through interactions with 13-mer sequences on one strand of initially unwound DNA [1] and with the subunits of DNA polymerase [2], the initiation protein facilitates strand specific replisome assembly at the plasmid replication origin. This step determines the direction of DNA replication. Interaction of replication initiator with β-subunit of DNA polymerase has been identified as an essential contribution of the plasmid replication initiation protein to the process of the replisome assembly. We propose models of the plas-mid replication initiation protein and its complex formed with single stranded DNA of replication origin. Our study provides new insights i...

Many factors leading to unfolding and misfolding of proteins eventually result in protein aggrega... more Many factors leading to unfolding and misfolding of proteins eventually result in protein aggregation. Stress imposed by high temperature was one of the first aggregation-inducing factors studied, and remains one of the main models in this field. The cell needs chaperone proteins to control and counteract the aggregation process. Elimination of aggregates can be achieved by solubilization of aggregates and either refolding of the liberated polypeptides or their proteolysis. Here we focus on the molecular mechanisms by which small Hsps chaperones and chaperones from Hsp100 family in cooperation with Hsp70 and Hsp40 chaperones liberate and refold polypeptides trapped in protein aggregates. It has been proposed that small Hsps associate with aggregating polypeptides, thus changing their biochemical properties so that the subsequent Hsp100-Hsp70 disaggregation process becomes much more efficient. Two members of the small Hsp family, IbpA and IbpB, are present in Escherichia coli. Their ...

Nucleic Acids Research, 2021

An essential feature of replication initiation proteins is their ability to bind to DNA. In this ... more An essential feature of replication initiation proteins is their ability to bind to DNA. In this work, we describe a new domain that contributes to a replication initiator sequence-specific interaction with DNA. Applying biochemical assays and structure prediction methods coupled with DNA–protein crosslinking, mass spectrometry, and construction and analysis of mutant proteins, we identified that the replication initiator of the broad host range plasmid RK2, in addition to two winged helix domains, contains a third DNA-binding domain. The phylogenetic analysis revealed that the composition of this unique domain is typical within the described TrfA-like protein family. Both in vitro and in vivo experiments involving the constructed TrfA mutant proteins showed that the newly identified domain is essential for the formation of the protein complex with DNA, contributes to the avidity for interaction with DNA, and the replication activity of the initiator. The analysis of mutant proteins...

Journal of Chemical Information and Modeling, 2020

The method for protein-structure prediction, which combines the physics-based coarse-grained UNRE... more The method for protein-structure prediction, which combines the physics-based coarse-grained UNRES force field with knowledge-based modeling, has been developed further and tested in the 13th Community Wide Experiment on the Critical Assessment of Techniques for Protein Structure Prediction (CASP13). The method implements restraints from the consensus fragments common to server models. In this work, the server models to derive fragments have been chosen on the basis of quality assessment; a fully automatic fragment-selection procedure has been introduced, and Dynamic Fragment Assembly pseudopotentials have been fully implemented. The Global Distance Test Score (GDT_TS), averaged over our "Model 1" predictions, increased by over 10 units with respect to CASP12 for the freemodeling category to reach 40.82. Our "Model 1" predictions ranked 20 and 14 for all and free-modeling targets, respectively (upper 20.2% and 14.3% of all models submitted to CASP13 in these categories, respectively), compared to 27 (upper 21.1%) and 24 (upper 18.9%) in CASP12, respectively. For oligomeric targets, the Interface Patch Similarity (IPS) and Interface Contact Similarity (ICS) averaged over our best oligomer models increased from 0.28 to 0.36 and from 12.4 to 17.8, respectively, from CASP12 to CASP13, and top-ranking models of 2 targets (H0968 and T0997o) were obtained (none in CASP12). The improvement of our method in CASP13 over CASP12 was ascribed to the combined effect of the overall enhancement of server-model quality, our success in selecting server models and fragments to derive restraints, and improvements of the restraint and potential-energy functions.

Journal of Molecular Graphics and Modelling, 2019

The recent NEWCT-9P version of the coarse-grained UNRES force field for proteins, with scale-cons... more The recent NEWCT-9P version of the coarse-grained UNRES force field for proteins, with scale-consistent formulas for the local and correlation terms, has been tested in the CASP13 experiment of the blind-prediction of protein structure, in the ab initio, contact-assisted, and data-assisted modes. Significant improvement of the performance has been observed with respect to the CASP11 and CASP12 experiments (by over 10 GDT_TS units for the ab initio mode predictions and by over 15 GDT_TS units for the contact-assisted prediction, respectively), which is a result of introducing scale-consistent terms and improved handling of contact-distance restraints. As in previous CASP exercises, UNRES ranked higher in the free modeling category than in the general category that included template based modeling targets. Use of distance restraints from the predicted contacts, albeit many of them were wrong, resulted in the increase of GDT_TS by over 8 units on average and introducing sparse restraints from small-angle X-ray/neutron scattering and chemical cross-link-mass-spectrometry experiments, and ambiguous restraints from nuclear magnetic resonance experiments has also improved the predictions by 8.6, 9.7, and 10.7 GDT_TS units on average, respectively.

Frontiers in Molecular Biosciences, 2016

The proper initiation and occurrence of DNA synthesis depends on the formation and rearrangements... more The proper initiation and occurrence of DNA synthesis depends on the formation and rearrangements of nucleoprotein complexes within the origin of DNA replication. In this review article, we present the current knowledge on the molecular mechanism of replication complex assembly at the origin of bacterial chromosome and plasmid replicon containing direct repeats (iterons) within the origin sequence. We describe recent findings on chromosomal and plasmid replication initiators, DnaA and Rep proteins, respectively, and their sequence-specific interactions with double-and single-stranded DNA. Also, we discuss the current understanding of the activities of DnaA and Rep proteins required for replisome assembly that is fundamental to the duplication and stability of genetic information in bacterial cells.

Org. Biomol. Chem., 2015

Introduction of hydroxyethylene isosteres into glycopeptides led to loss of Aq affinity and subse... more Introduction of hydroxyethylene isosteres into glycopeptides led to loss of Aq affinity and subsequent T cell response due to disruption of hydrogen bond networks.

Methods and Principles in Medicinal Chemistry, 2010

Methods and Principles in Medicinal Chemistry, 2011

ABSTRACT

MedChemComm, 2012

Novel thiobarbiturates were identified by virtual screening and MM-PBSA calculation as potent sir... more Novel thiobarbiturates were identified by virtual screening and MM-PBSA calculation as potent sirtuin inhibitors which represent useful probes for cellular studies.

Journal of Medicinal Chemistry, 2012

The diphtheria toxin-like ADP-ribosyltransferases (ARTDs) are an enzyme family that catalyses the... more The diphtheria toxin-like ADP-ribosyltransferases (ARTDs) are an enzyme family that catalyses the transfer of ADP-ribose units onto substrate proteins, using nicotinamide adenine dinucleotide (NAD +) as a co-substrate. They have a documented role in chromatin remodelling and DNA repair; and inhibitors of ARTD1 and 2 (PARP1 and 2) are currently in clinical trials for the treatment of cancer. The detailed function of most other ARTDs is still unknown. Using virtual screening we identified small ligands of ARTD7 (PARP15/BAL3) and ARTD8 (PARP14/BAL2). Thermal-shift assays confirmed that 16 compounds, belonging to eight structural classes, bound to ARTD7/ARTD8. Affinity measurements with isothermal titration calorimetry for two isomers of the most promising hit compound confirmed binding in the low micromolar range to ARTD8. Crystal structures showed anchoring of the hits in the nicotinamide pocket. These results form a starting point in the development of chemical tools for the study of the role and function of ARTD7 and ARTD8.

Journal of Medicinal Chemistry, 2010

Class III histone deacetylases (sirtuins) play pivotal roles in many cellular processes. They are... more Class III histone deacetylases (sirtuins) play pivotal roles in many cellular processes. They are linked to extended lifespan and to the pathogenesis of cancer and neuronal disorders. We present novel sirtuin inhibitors based on a 6,7-dichloro-2-oxindole scaffold with low micromolar activity. In vitro activity was rationalized by docking studies, and hyperacetylation of sirtuin targets could be demonstrated in cell culture.

Journal of Medicinal Chemistry, 2011

Parts of this thesis have already been published in scientific journals, and presented at scienti... more Parts of this thesis have already been published in scientific journals, and presented at scientific conferences.

ChemMedChem, 2008

NAD+-dependent histone deacetylases (sirtuins) are enzymes that cleave acetyl groups from lysine ... more NAD+-dependent histone deacetylases (sirtuins) are enzymes that cleave acetyl groups from lysine residues in histones and other proteins. Potent selective sirtuin inhibitors are interesting tools for the investigation of the biological functions of these enzymes and may be future drugs for the treatment of cancer or neurodegenerative diseases. Herein we present the results from a protein-based virtual screen of a commercial database with subsequent biological testing of the most promising compounds. The combination of docking and in vitro experimental testing resulted in the identification of novel sirtuin inhibitors with thiobarbiturate structure. To rationalize the experimental results, free-energy calculations were carried out by molecular mechanics Poisson-Boltzmann/surface area (MM-PBSA) calculations. A significant correlation between calculated binding free energies and measured Sirt2 inhibitory activities was observed. The analyses suggested a molecular basis for the interaction of the identified thiobarbiturate derivatives with human Sirt2. Based on the docking and MM-PBSA calculations we synthesized and tested five further thiobarbiturates. The MM-PBSA method correctly predicted the activity of the novel thiobarbiturates. The identified compounds will be used to further explore the therapeutic potential of sirtuin inhibitors.

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, 2010

Histone deacetylases (HDACs) are enzymes that cleave acetyl groups from acetyl-lysine residues in... more Histone deacetylases (HDACs) are enzymes that cleave acetyl groups from acetyl-lysine residues in histones and various nonhistone proteins. Unlike the other three of the four classes of HDACs that have been identified in humans, which are zinc-dependent amidohydrolases, class III HDACs depend on nicotinamide adenine dinucleotide (NAD +) for their catalytic activity. The seven members of the class III HDACs are also named sirtuins for their homology to Sir2p, a yeast histone deacetylase. Sirtuin inhibitors have been critical for the linkage of sirtuin activity to many physiological and pathological processes, and sirtuin activity has been associated with the pathogenesis of cancer, HIV, and metabolic and neurological diseases. Presented here is an overview of the many sirtuin inhibitors that have provided insight into the biological role of sirtuins.

Medicinal Research Reviews, 2009

Histone deacetylases (HDACs) are enzymes that cleave off acetyl groups from acetyl-lysine residue... more Histone deacetylases (HDACs) are enzymes that cleave off acetyl groups from acetyl-lysine residues in histones and various nonhistone proteins. Four different classes of HDACs have been identified in humans so far. Although classes I, II, and IV are zinc-dependent amidohydrolases, class III HDACs depend on nicotinamide adenine dinucleotide (NAD 1) for their catalytic activity. According to their homology to Sir2p, a yeast histone deacetylase, the class III is also termed sirtuins. Seven members have been described in humans so far. As sirtuins are involved in many physiological and pathological processes, their activity has been associated with the pathogenesis of cancer, HIV, metabolic, or neurological diseases. Herein, we present an overview over sirtuins including their biology, targets, inhibitors, and activators and their potential as new therapeutic agents.