Interakce Proteinů s Nukleovými Kyselinami: od Struktury k Specificitě

Sequence-specific interactions between proteins and nucleic acids play an essential role in the cell biology. While several molecular mechanisms contributing to the binding speci- ficity have been identified empirically, no general protein-DNA recognition code has been described to date. In this thesis, I explore selected characteristics of protein-DNA inter- actions using computational methods. First, the pairwise interactions between the basic biomolecular building blocks-amino acids and nucleotides-are investigated. It is shown that several statistically enriched, biologically relevant interaction motifs correspond to the most energetically favorable configurations of the respective binding partners. In ad- dition, a relationship between the physico-chemical properties of the amino acid residues found at the protein-DNA interface and the local geometric features of the DNA helix is presented. Next, the applicability of molecular dynamics-based setups to the description of binding equilibria in protein-DNA systems is investigated. Discrepancies are observed between the description offered by the computer simulations and experimental results, as well as between the results obtained using two molecular mechanical force fields. Finally, the more general evolutionary aspects of protein organization...Sekvenčně-specifické interakce mezi proteiny a nukleovými kyselinami mají zásadní roli v biologii buňky. I když několik molekulárních mechanismů podílejících se na vazebné speci- ficitě bylo empiricky vypozorováno, žádný obecný rozpoznávací kód pro protein-DNA interakce nebyl zatím popsán. V této disertační práci prozkoumávám vybrané charakter- istiky protein-DNA interakcí pomocí výpočetních metod. Nejdřív jsou studovány párové interakce mezi základními stavebními prvky biomolekul-aminokyselinami a nukleotidy. Je ukázáno, že některé statisticky nabohacené, biologicky relevantní interakční motivy odpovídají energeticky nejvýhodnějším geometrickým uspořádáním daných vazebných partnerů. Dále je demonstrován vztah mezi fyzikálně-chemickými vlastnostmi aminoky- selin nacházejících se na rozhraní proteinu s DNA a lokálními topologickými charak- teristikami DNA dvoušroubovice. V další části je prozkoumána využitelnost postupů založených na molekulové dynamice při popisu vazebných rovnováh v systémech protein- DNA. Jsou pozorovány rozdíly nejen mezi popisem získaným na základě počítačových simulací a experimentálními výsledky, ale i mezi výsledky získanými s využitím dvou různých molekulárně mechanických silových polí. V závěru jsou prozkoumány obecnější evoluční charakteristiky struktury proteinů a je...Department of BiochemistryKatedra biochemiePřírodovědecká fakultaFaculty of Scienc

Metabolismus aminokyselin u parazitických a anaerobních protist

Parazitické prvoky sú príčinou nespočetných patologických stavov a ekonomických problémov v mnohých častiach sveta. Napriek ich fylogenetickej nepríbuznosti u nich nachádzame spoločné rysy v ich prístupe k uspokojovaniu základných životných potrieb. Na rozdiel od študovanejšieho energetického metabolizmu, dráhy využitia amino kyselín sú v mnohých oblastiach neznáme. Prehľad ukazuje, že parazitický životný štyl na metabolizmus amino kyselín nemal až taký závažný dopad ako na energetický metabolizmus, ktorý je často výrazne zjednodušený. Objavili sa u nich nové dráhy biosyntézy amino kyselín tak esenciálnych, ako aj neesenciálnych u ľudí. Arginín dihydrolázová dráha u Trichomonas a Giardia predstavuje kompletne nový spôsob využitia danej amino kyseliny. Metabolizmus síru-obsahujúcich amino kyselín je predmetom intenzívnejšieho výskumu kvôli ich roliam mimo proteogenézu. Polyamíny sú dusíkaté organické látky hrajúce úlohu v mnohých procesoch v bunke, vrátane replikácie DNA a translácie proteínov. Objasnená je syntéza polyamínov a ich derivátov, ktorá ja na metabolizmus amino kyselín priamo napojená. Zrhnutie metód využívajúcich unikátne dráhy nájdené u parazitických prvokov v sebe spája predchádzajúci výskum so zavedením amino kyselinových metabolizmov ako zatiaľ nevyužitých cieľov liečiv.Parasitic protists are the cause of countless pathological conditions and economic issues in many parts of the world. While being phylogenetically unrelated, they share many similarities in their approach to satisfying the essential needs. Unlike the much studied energy metabolism, amino acids utilisation pathways are rather unexplored areas. This review shows that in many cases, the parasitic life style has not had the same impact on the amino acid metabolism as it did on the energy metabolism of the protists, which is often severely reduced. Novel pathways have been found in many of the organisms in question, for the biosynthesis of amino acids deemed both essential and non-essential in humans. The arginine dihydrolase pathway found in Trichomonas and Giardia represents a complely new way of utilising the said amino acid. The metabolism of sulfur-containing amino acid has been a matter of intensive research for their non-proteogenic roles. Polyamines are organic nitrogenous compounds involved in many vital processes in the cells, including DNA replication and protein translation. The synthesis of polyamines and their derivatives is elucidated, as it is directly connected to the amino acid metabolism. Finally, the exploitation of the unique pathways described integrates the previous research with the aim...Department of ParasitologyKatedra parazitologieFaculty of SciencePřírodovědecká fakult

Interakční preference v komplexech protein - DNA.

Interakční preference v komplexes protein - DNA Dávid Jakubec Abstrakt Interakce proteinů s DNA jsou základem mnoha esenciálních biologických pochodů. Navzdory dosavadním snahám se zatím nepodařilo kompletně objasnit pravidla řídící rozpoznávání specifických úseků nukleových kyselin proteiny. V této práci se pokouším prozkoumat proces rozpoznávání DNA rozdělením složité sítě kontaktů na rozhraní protein - DNA do příspěvků jednotlivých párů aminokyselina - nukleotid. Tyto páry byly získány z exis- tujících struktur protein - DNA komplexů ve vysokém rozlišení a zpracovány bioinformatickými metodami a nástroji výpočetné chemie. Nově jsem zavedl kritéria specificity sprahující pozorované geometrické preference s relativní energetickou bilancí párů. Aplikací těchto kritérií jsem rozšířil knihovnu párů aminokyselina - nukleotid které se mohou podílet na přímém rozpoznávání sekvence. S cílem prozkoumat fyzikální základy pozorované specificity jsem vypočítal mapy elektrostatických potenciálů pro jednotlivé nukleotidy a vy- brané komplexy. 1Interaction preferences in protein - DNA complexes Dávid Jakubec Abstract Interactions of proteins with DNA lie at the basis of many fundamental bio- logical processes. Despite ongoing efforts, the rules governing the recognition of specific nucleic acid sequences have still not been universally elucidated. In this work, I attempt to explore the recognition process by splitting the intricate network of contacts at the protein - DNA interface into contribu- tions of individual amino acid - nucleotide pairs. These pairs are extracted from existing high-resolution structures of protein - DNA complexes and in- vestigated by bioinformatics and computational-chemistry based methods. Criteria of specificity based on the coupling of observed geometrical prefer- ences and the respective interaction energies are introduced. The application of these criteria is used to expand the library of amino acid - nucleotide pairs potentially significant for direct sequence recognition. Electrostatic poten- tial maps are calculated for individual nucleotides as well as for selected complexes to investigate the physical basis of the observed specificity. 1Department of BiochemistryKatedra biochemiePřírodovědecká fakultaFaculty of Scienc

Interactions of Proteins with Nucleic Acids: from Structure to Specificity

Sequence-specific interactions between proteins and nucleic acids play an essential role in the cell biology. While several molecular mechanisms contributing to the binding speci- ficity have been identified empirically, no general protein-DNA recognition code has been described to date. In this thesis, I explore selected characteristics of protein-DNA inter- actions using computational methods. First, the pairwise interactions between the basic biomolecular building blocks-amino acids and nucleotides-are investigated. It is shown that several statistically enriched, biologically relevant interaction motifs correspond to the most energetically favorable configurations of the respective binding partners. In ad- dition, a relationship between the physico-chemical properties of the amino acid residues found at the protein-DNA interface and the local geometric features of the DNA helix is presented. Next, the applicability of molecular dynamics-based setups to the description of binding equilibria in protein-DNA systems is investigated. Discrepancies are observed between the description offered by the computer simulations and experimental results, as well as between the results obtained using two molecular mechanical force fields. Finally, the more general evolutionary aspects of protein organization..

Interaction preferences in protein - DNA complexes

Interaction preferences in protein - DNA complexes Dávid Jakubec Abstract Interactions of proteins with DNA lie at the basis of many fundamental bio- logical processes. Despite ongoing efforts, the rules governing the recognition of specific nucleic acid sequences have still not been universally elucidated. In this work, I attempt to explore the recognition process by splitting the intricate network of contacts at the protein - DNA interface into contribu- tions of individual amino acid - nucleotide pairs. These pairs are extracted from existing high-resolution structures of protein - DNA complexes and in- vestigated by bioinformatics and computational-chemistry based methods. Criteria of specificity based on the coupling of observed geometrical prefer- ences and the respective interaction energies are introduced. The application of these criteria is used to expand the library of amino acid - nucleotide pairs potentially significant for direct sequence recognition. Electrostatic poten- tial maps are calculated for individual nucleotides as well as for selected complexes to investigate the physical basis of the observed specificity.

Interaction preferences in protein - DNA complexes

Interaction preferences in protein - DNA complexes Dávid Jakubec Abstract Interactions of proteins with DNA lie at the basis of many fundamental bio- logical processes. Despite ongoing efforts, the rules governing the recognition of specific nucleic acid sequences have still not been universally elucidated. In this work, I attempt to explore the recognition process by splitting the intricate network of contacts at the protein - DNA interface into contribu- tions of individual amino acid - nucleotide pairs. These pairs are extracted from existing high-resolution structures of protein - DNA complexes and in- vestigated by bioinformatics and computational-chemistry based methods. Criteria of specificity based on the coupling of observed geometrical prefer- ences and the respective interaction energies are introduced. The application of these criteria is used to expand the library of amino acid - nucleotide pairs potentially significant for direct sequence recognition. Electrostatic poten- tial maps are calculated for individual nucleotides as well as for selected complexes to investigate the physical basis of the observed specificity.

The metabolism of amino acids in parasitic and anaerobic protists

Parasitic protists are the cause of countless pathological conditions and economic issues in many parts of the world. While being phylogenetically unrelated, they share many similarities in their approach to satisfying the essential needs. Unlike the much studied energy metabolism, amino acids utilisation pathways are rather unexplored areas. This review shows that in many cases, the parasitic life style has not had the same impact on the amino acid metabolism as it did on the energy metabolism of the protists, which is often severely reduced. Novel pathways have been found in many of the organisms in question, for the biosynthesis of amino acids deemed both essential and non-essential in humans. The arginine dihydrolase pathway found in Trichomonas and Giardia represents a complely new way of utilising the said amino acid. The metabolism of sulfur-containing amino acid has been a matter of intensive research for their non-proteogenic roles. Polyamines are organic nitrogenous compounds involved in many vital processes in the cells, including DNA replication and protein translation. The synthesis of polyamines and their derivatives is elucidated, as it is directly connected to the amino acid metabolism. Finally, the exploitation of the unique pathways described integrates the previous research with the aim..