Zastosowanie biofizyki obliczeniowej i bioinformatyki w wielkoskalowych problemach biologicznych

In my thesis I presented two applications of bioinformatics and computational biophysics in multiscale biological problems. The first project aimed to provide insights into enzymatic methylation in S. cerevisiae using the bioinformatic approach. I conducted a comprehensive structural and functional classification of all the methyltransferases encoded in its genome. Using distant homology detection and fold recognition, I found that the S. cerevisiae methyltransferome comprises 86 MTases (53 well-known and 33 putative with unknown substrate specificity). Structural classification of their catalytic domains shows that these enzymes may adopt nine different folds, the most common being the Rossmann-like. I also analyzed the domain architecture of these proteins and identified several new domain contexts. I found that the majority of MTase genes are periodically expressed during yeast metabolic cycle. I used this finding, together with the calculated isoelectric point, fold assignment and cellular localization, to develop a novel method for predicting substrate specificity. Moreover, I predicted the general substrate for 24 of 33 putative MTases using this approach. My predictions were correct for eight proteins out of twelve putative MTases already experimentally verified. What is important, my methodology can be further extended to the other protein families. Finally, I showed that, in S. cerevisiae, methylation is carried out by 35 RNA MTases, 31 protein MTases, nine small molecule MTases, three lipid MTases and eight MTases with unknown substrate specificity. In the second project I applied the computational biophysics approach to describe protein-protein interactions in ubiquitin recognition. I performed an extensive structural analysis of a large set of bound and unbound ubiquitin conformers, and studied the level of residual induced fit after conformational selection in the binding process. I showed that the region surrounding the binding site in ubiquitin undergoes conformational changes that are significantly more pronounced compared with the whole molecule on the average. I demonstrated that these induced fit structural adjustments are comparable in magnitude to the conformational selection. Moreover, I demonstrated that this picture is preserved in other structural ensembles based on the same experimental data. Finally I proposed a model of ubiquitin binding, which blends conformational selection with subsequent induced fit, and provides a quantitative measure of their respective contributions.Moja praca doktorska przedstawia zastosowania bioinformatyki oraz biofizyki obliczeniowej w wieloskalowych problemach biologicznych. W pierwszej części doktoratu zastosowałem narzędzia bioinformatyki w badaniu metylacji u S. cerevisiae. W obrębie tej pracy przeprowadziłem obszerną klasyfikację, pod kątem struktury oraz funkcji, wszystkich metylotransferaz (MTaz) kodowanych w genomie tego drożdża. Wykazałem, stosując metody detekcji dalekiej homologii, iż metylotransferom S. cerevisiae składa się z 86 MTaz, pośród nich 53 MTazy zostały już wcześniej eksperymentalnie scharakteryzowane, natomiast dla pozostałych 33 wciąż nie mamy potwierdzonej aktywności metylotransferazowej. Przeprowadzona przeze mnie klasyfikacja strukturalna domeny katalitycznej pokazała, iż metylotransferazy mogą przyjmować 9 różnych zwojów, przy czym dominującym jest zwój Rossmann-like. Ponadto przeanalizowałem architekturę domenową tych białek i zidentyfikowałem kilka nowych kontekstów domenowych. W swojej pracy wskazałem również, iż większość genów MTaz podlega cyklicznej ekspresji podczas drożdżowego cyklu metabolicznego (yeast metabolic cycle). Wykorzystałem tę obserwację, uzupełniając ją danymi o punkcie izoelektrycznym, rodzaju zwoju i lokalizacji komórkowej, do zaproponowania nowej metody przewidywania specyficzności substratowej. Metoda ta, działająca w oparciu o drzewa decyzyjne, pozwoliła mi na przewidzenie specyficzności substratowej dla 24 z 33 potencjalnych MTaz. Moje przewidywania zostały już potwierdzone dla ośmiu z dwunastu do tej pory zweryfikowanych eksperymentalnie nowych MTaz. Ponadto zaproponowany przeze mnie algorytm może być zastosowany do badania kolejnych rodzin białkowych. Podsumowując, wykazałem iż u S. cerevisiae metylacja jest przeprowadzana przez 35 MTaz RNA, 31 MTaz białkowych, dziewięć MTaz małych cząsteczek, trzy MTazy lipidów a także osiem MTaz, dla których wciąż nie znamy specyficzności substratowej. W drugiej części doktoratu zastosowałem narzędzia biofizyki obliczeniowej do opisu oddziaływania białko-białko w przypadku ubikwityny. Przeprowadziłem dokładną analizę strukturalną zbioru konformacji związanej jak i wolnej ubikwityny oraz zbadałem zakres stosowalności modelu wymuszonego dopasowania (induced fit), następującego po selekcji konformacyjnej (conformational selection), w opisie procesu wiązania ubikwityny. W mojej pracy wykazałem, iż w otoczeniu miejsca wiązania liganda do ubikwityny zachodzą zmiany konformacyjne, które są większe w porównaniu do średnich zmian strukturalnych w obrębie całego białka. Następnie pokazałem, iż te lokalne zmiany wyjaśnia model wymuszonego dopasowania a ich zakres ich jest porównywalna do zmian wywołanych selekcją konformacyjną. Podsumowując, w mojej pracy zaproponowałem nowy model opisujący oddziaływanie ubikwityny z ligandami białkowymi, model ten łączy selekcje konformacyjną z następującym po niej wymuszonym dopasowaniem zachodzącym w obrębie miejsca wiązania. Ponadto przedstawiłem nowy sposób badania udziału wymuszonego dopasowania i selekcji konformacyjne w opisie oddziaływania białko-białko

Structural and Optical Characterization of ZnS Ultrathin Films Prepared by Low-Temperature ALD from Diethylzinc and 1.5-Pentanedithiol after Various Annealing Treatments

The structural and optical evolution of the ZnS thin films prepared by atomic layer deposition (ALD) from the diethylzinc (DEZ) and 1,5-pentanedithiol (PDT) as zinc and sulfur precursors was studied. A deposited ZnS layer (of about 60 nm) is amorphous, with a significant S excess. After annealing, the stoichiometry improved for annealing temperatures ≥400 °C and annealing time ≥2 h, and 1:1 stoichiometry was obtained when annealed at 500 °C for 4 h. ZnS crystallized into small crystallites (1–7 nm) with cubic sphalerite structure, which remained stable under the applied annealing conditions. The size of the crystallites (D) tended to decrease with annealing temperature, in agreement with the EDS data (decreased content of both S and Zn with annealing temperature); the D for samples annealed at 600 °C (for the time ≤2 h) was always the smallest. Both reflectivity and ellipsometric spectra showed characteristics typical for quantum confinement (distinct dips/peaks in UV spectral region). It can thus be concluded that the amorphous ZnS layer obtained at a relatively low temperature (150 °C) from organic S precursor transformed into the layers built of small ZnS nanocrystals of cubic structure after annealing at a temperature range of 300–600 °C under Ar atmosphere

Spatial variability of selected physicochemical parameters within peat deposits in small valley mire: a geostatistical approach

Geostatistical methods for 2D and 3D modelling spatial variability of selected physicochemical properties of biogenic sediments were applied to a small valley mire in order to identify the processes that lead to the formation of various types of peat. A sequential Gaussian simulation was performed to reproduce the statistical distribution of the input data (pH and organic matter) and their semivariances, as well as to honouring of data values, yielding more ‘realistic’ models that show microscale spatial variability, despite the fact that the input sample cores were sparsely distributed in the X–Y space of the study area. The stratigraphy of peat deposits in the Ldzań mire shows a record of long-term evolution of water conditions, which is associated with the variability in water supply over time. Ldzań is a fen (a rheotrophic mire) with a through-flow of groundwater. Additionally, the vicinity of the Grabia River is marked by seasonal inundations of the southwest part of the mire and increased participation of mineral matter in the peat. In turn, the upper peat layers of some of the central part of Ldzań mire are rather spongy, and these peat-forming phytocoenoses probably formed during permanent waterlogging

Structural and Optical Characterization of ZnS Ultrathin Films Prepared by Low-Temperature ALD from Diethylzinc and 1.5-Pentanedithiol after Various Annealing Treatments

The structural and optical evolution of the ZnS thin films prepared by atomic layer deposition (ALD) from the diethylzinc (DEZ) and 1,5-pentanedithiol (PDT) as zinc and sulfur precursors was studied. A deposited ZnS layer (of about 60 nm) is amorphous, with a significant S excess. After annealing, the stoichiometry improved for annealing temperatures ≥400 °C and annealing time ≥2 h, and 1:1 stoichiometry was obtained when annealed at 500 °C for 4 h. ZnS crystallized into small crystallites (1–7 nm) with cubic sphalerite structure, which remained stable under the applied annealing conditions. The size of the crystallites (D) tended to decrease with annealing temperature, in agreement with the EDS data (decreased content of both S and Zn with annealing temperature); the D for samples annealed at 600 °C (for the time ≤2 h) was always the smallest. Both reflectivity and ellipsometric spectra showed characteristics typical for quantum confinement (distinct dips/peaks in UV spectral region). It can thus be concluded that the amorphous ZnS layer obtained at a relatively low temperature (150 °C) from organic S precursor transformed into the layers built of small ZnS nanocrystals of cubic structure after annealing at a temperature range of 300–600 °C under Ar atmosphere

Pituitary tumours — a large retrospective single-centre study of over 2300 cases. Experience of a tertiary reference centre

Introduction: Pituitary adenomas (PAs), also known as a pituitary neuroendocrine tumours (PitNET), are usually benign tumours of the anterior lobe of the pituitary gland and account for the third most common intracranial neoplasm. The most common type of pituitary adenoma is lactotroph adenoma, in which dopamine agonists are the first-line treatment. Nevertheless, in selected cases surgery or even radiotherapy may be required. In the current study, we aimed to analyse all patients who underwent surgery due to intrasellar mass in order to evaluate frequency of particular pituitary tumours, clinical diagnosis, and pathology findings. Material and methods: We retrospectively analysed all cases of patients consecutively operated due to intrasellar mass between 1st January 2010 and 31st December 2018 at the Department of Neurosurgery, Military Institute of Medicine in Warsaw, Poland. Results: Our database included 2348 cases: 1390 women (59.2%) and 958 men (40.8%). The mean age for women was 48.4 years (SD ± 15.72; median 49) and for men 50.9 years (SD ± 14.94; median 53). In our cohort we found: 869 gonadotroph and null cell adenomas, 751 somatotroph and mammosomatotroph adenomas, 386 corticotroph adenomas, 71 plurihormonal adenomas, 59 craniopharyngiomas, 44 lactotroph adenomas, 18 purely thyrotroph adenomas, and other rare cases of pituitary tumours including one pituitary carcinoma metastasising to the liver (corticotroph origin). Conclusions: We provide a comprehensive analysis of both clinical and pathological findings of the largest cohort of patients operated on for pituitary adenomas in one tertiary reference centre. To the best of our knowledge, this is the largest up-to-date published analysis in our country.

The ribosome stabilizes partially folded intermediates of a nascent multi-domain protein

Co-translational folding is crucial to ensure the production of biologically active proteins. The ribosome can alter the folding pathways of nascent polypeptide chains, yet a structural understanding remains largely inaccessible experimentally. We have developed site-specific labelling of nascent chains to detect and measure, using 19F nuclear magnetic resonance (NMR) spectroscopy, multiple states accessed by an immunoglobulin-like domain within a tandem repeat protein during biosynthesis. By examining ribosomes arrested at different stages during translation of this common structural motif, we observe highly broadened NMR resonances attributable to two previously unidentified intermediates, which are stably populated across a wide folding transition. Using molecular dynamics simulations and corroborated by cryo-electron microscopy, we obtain models of these partially folded states, enabling experimental verification of a ribosome-binding site that contributes to their high stabilities. We thus demonstrate a mechanism by which the ribosome could thermodynamically regulate folding and other co-translational processes

Communicative Planes of Discourse Revisited: Language, Text, Register and Genre

Communicative Planes of Discourse Revisited: Language, Text, Register and GenreThe article raises the issue of correlation between various interconnected facets of discourse, such as language, text, register and genre. Though these notions are well entrenched within discourse studies, it is difficult to offer clear-cut definitions of them, because the phenomena to which they refer are extremely complex and multi-faceted. This article constitutes one of numerous endeavours to clarify these notions and the correlations among them. At the outset, the article defines two of the most basic and, thus, indispensable notions for any discourse analysis, i.e. the notions of language and text, which are herein described in cognitive terms as symbolic assemblies Evans and Green 2006. Language and text differ from each other with respect to the nature of the semiotic plane they constitute: language is the plane of resources for verbal communication see Martin 1992, whilst texts are the plane of the manifestation and realisation of such resources. Next, the article clarifies that the realisation of language in particular texts is mediated by the semiotic planes of register and genre, the former concerning the situational variety of the language, and the latter concerning conventional patterns of whole texts Biber and Conrad 2009. Both register and genre are herein regarded in terms of structures of knowledge concerning WHAT and HOW to communicate in given situational and contextual circumstances; hence, they are regarded as abstract semiotic planes through which language is manifested in particular texts/utterances. Thus, the article advocates the stance that the levels of register and genre constitute the plane of mediation, whilst the level of concrete texts/utterances constitutes the plane of the manifestation and realisation of language in its registers and genres. The article refers not only to studies within the English language but also to relevant areas of Polish genre studies.Communicative Planes of Discourse Revisited: Language, Text, Register and GenreThe article raises the issue of correlation between various interconnected facets of discourse, such as language, text, register and genre. Though these notions are well entrenched within discourse studies, it is difficult to offer clear-cut definitions of them, because the phenomena to which they refer are extremely complex and multi-faceted. This article constitutes one of numerous endeavours to clarify these notions and the correlations among them. At the outset, the article defines two of the most basic and, thus, indispensable notions for any discourse analysis, i.e. the notions of language and text, which are herein described in cognitive terms as symbolic assemblies Evans and Green 2006. Language and text differ from each other with respect to the nature of the semiotic plane they constitute: language is the plane of resources for verbal communication see Martin 1992, whilst texts are the plane of the manifestation and realisation of such resources. Next, the article clarifies that the realisation of language in particular texts is mediated by the semiotic planes of register and genre, the former concerning the situational variety of the language, and the latter concerning conventional patterns of whole texts Biber and Conrad 2009. Both register and genre are herein regarded in terms of structures of knowledge concerning WHAT and HOW to communicate in given situational and contextual circumstances; hence, they are regarded as abstract semiotic planes through which language is manifested in particular texts/utterances. Thus, the article advocates the stance that the levels of register and genre constitute the plane of mediation, whilst the level of concrete texts/utterances constitutes the plane of the manifestation and realisation of language in its registers and genres. The article refers not only to studies within the English language but also to relevant areas of Polish genre studies