From DNA sequences to chemical structures – methods for mining microbial genomic and metagenomic data sets for new natural products

Rapid mining of large genomic and metagenomic data sets for modular polyketide synthases, non-ribosomal peptide synthetases and hybrid polyketide synthase/non-ribosomal peptide synthetase biosynthetic gene clusters has been achieved using the generic computer program packages ClustScan and CompGen. These program packages perform the annotation with the hierarchical structuring into polypeptides, modules and domains, as well as storage and graphical presentations of the data. This aims to achieve the most accurate predictions of the activities and specificities of catalytically active domains that can be made with present knowledge, leading to a prediction of the most likely chemical structures produced by these enzymes. The program packages also allow generation of novel clusters by homologous recombination of the annotated genes in silico. ClustScan and CompGen were used to construct a custom database of known compounds (CSDB) and of predicted entirely novel recombinant products (r-CSDB) that can be used for in silico screening with computer-aided drug design technology. The use of these programs has been exemplified by analysing genomic sequences from terrestrial prokaryotes and eukaryotic microorganisms, a marine metagenomic data set and a newly discovered example of a 'shared metabolic pathway' in marine-microbial endosymbiosis

Nutrigenomics analysis of metabolic pathways in Saccharomyces cerevisiae followed by reading of reverse translation via De novo peptide sequencing

U ovome radu je razvijen novi ekspertni računalni sustav za analizu spektara masa putem kojeg je moguće tehnikom reverznog iščitavanja translacije, odnosno identifikacije proteina u uvjetima de novo, utvrditi promjene na razini proteoma i posredno metaboloma u modelnom organizmu kvascu S. cerevisiae, čiji je genom u potpunosti sekvenciran. Ekspertni sustav razvijen je u Java programskom jeziku te se sastoji od dva djela, jedan koji izvršava analize na strani poslužitelja i drugi koji ima upravljačku funkciju, a nalazi se na strani korisnika. De novo sekvenciranjem peptida možemo identificirati nadeksprimirane peptide (putem spektrometra masa MALDI-TOF/TOF), odnosno proteine. Na temelju identificiranih proteina napravljena je nutrigenomička analiza iščitavanjem reverzne translacije, odnosno određeno je uključivanje i isključivanje pojedinih metaboličkih putova s poveznicom na promjene nastale u proteomu, kod uzgoja kvasca u aerobnim i anaerobnim uvjetima te pri uskraćivanju i obogaćivanju prihrane kvasca. Utvrđeno je da se nadeksprimirani proteini stanice kvasca razlikuju ovisno o načinu uzgoja, čime se može zaključiti da se ekspresija gena te sinteza proteina, a time i aktivnost metaboličkih putova znatno mijenja pod stresnim promjenama prihrane stanica kvasca.As a result of this thesis, expert system for the management and analysis of MALDI-TOF/TOF generated spectra is developed. This bioinformatics tool serves as a support for the derivatisation and analysis of proteoms of any organism by the use of mass spectrometry. The expert system consists of the proprietary database containing gene and protein data from all publicly available resources with their related taxonomic divisions and sub-divisions. Other inovative part of this system is a graphical user interface (client) used for both input and management of data generated from the spectra analyses. All spectra analyses are carried out on the remote server using algortihms and solutions developed during the course of work on this thesis. As a biological model for testing efficiency for the derivatisation and proteome analysis, the proteome of Saccharomyces cerevisiae was used. S. cerevisiae cells were grown in both aerobic and anaerobic conditions with respective food deprivation as well as optimal food content. Finally, proteins were identified which are with their over or under expression responsible for metabolic changes in yeast under the growth in different environmental conditions

Development of computer program for the acquisition, processing, storage and raphical presentation of biological data

U tijeku je razvoj integralnog računalnog programskog paketa, CompGen, za modeliranje procesa homologne rekombinacije između poliketidnih/peptidnih genskih nakupina u uvjetima in silico. Okosnica će programskog paketa CompGen biti specifično strukturirana baza podataka koja povezuje biosintetski put sinteze sa sekvencijama DNA genskih nakupina. U ovom je radu strukturirana specifična baza podataka poliketidnih/ peptidnih genskih nakupina po uzoru na BioSQL. Razvijeni su i računalni programi za: prikupljanje i strukturiranje bioloških podataka; detaljan opis ("anotaciju") modula, domena i njihovih poveznica; anotaciju biosintetskih putova i grafičko prikazivanje dvodimenzionalnih i trodimenzionalnih kemijskih struktura linearnih lanaca poliketida/peptida objektno orijentiranim računalnim jezikom Java i skriptnim računalnim jezikom JavaScript.An integral generic program package, CompGen, is under development to model the process of homologous recombination between polyketide/peptide gene-clusters in silico. The heart of CompGen will be a specially structured database that will connect the biosynthetic order to the DNA sequences. In this work the specific database was structured based on BioSQL. The computer programs for the acquisition and structuring of biological data; the annotation of modules, domains and their dockers and linkers; the annotation of biosynthetic order and for the graphical presentation of two-dimensional and three-dimensional chemical structures of linear polyketide/peptide chains were developed using computer languages Java and JavaScript

Development of computer program for the acquisition, processing, storage and raphical presentation of biological data

U tijeku je razvoj integralnog računalnog programskog paketa, CompGen, za modeliranje procesa homologne rekombinacije između poliketidnih/peptidnih genskih nakupina u uvjetima in silico. Okosnica će programskog paketa CompGen biti specifično strukturirana baza podataka koja povezuje biosintetski put sinteze sa sekvencijama DNA genskih nakupina. U ovom je radu strukturirana specifična baza podataka poliketidnih/ peptidnih genskih nakupina po uzoru na BioSQL. Razvijeni su i računalni programi za: prikupljanje i strukturiranje bioloških podataka; detaljan opis ("anotaciju") modula, domena i njihovih poveznica; anotaciju biosintetskih putova i grafičko prikazivanje dvodimenzionalnih i trodimenzionalnih kemijskih struktura linearnih lanaca poliketida/peptida objektno orijentiranim računalnim jezikom Java i skriptnim računalnim jezikom JavaScript.An integral generic program package, CompGen, is under development to model the process of homologous recombination between polyketide/peptide gene-clusters in silico. The heart of CompGen will be a specially structured database that will connect the biosynthetic order to the DNA sequences. In this work the specific database was structured based on BioSQL. The computer programs for the acquisition and structuring of biological data; the annotation of modules, domains and their dockers and linkers; the annotation of biosynthetic order and for the graphical presentation of two-dimensional and three-dimensional chemical structures of linear polyketide/peptide chains were developed using computer languages Java and JavaScript

Nutrigenomics analysis of metabolic pathways in Saccharomyces cerevisiae followed by reading of reverse translation via De novo peptide sequencing

U ovome radu je razvijen novi ekspertni računalni sustav za analizu spektara masa putem kojeg je moguće tehnikom reverznog iščitavanja translacije, odnosno identifikacije proteina u uvjetima de novo, utvrditi promjene na razini proteoma i posredno metaboloma u modelnom organizmu kvascu S. cerevisiae, čiji je genom u potpunosti sekvenciran. Ekspertni sustav razvijen je u Java programskom jeziku te se sastoji od dva djela, jedan koji izvršava analize na strani poslužitelja i drugi koji ima upravljačku funkciju, a nalazi se na strani korisnika. De novo sekvenciranjem peptida možemo identificirati nadeksprimirane peptide (putem spektrometra masa MALDI-TOF/TOF), odnosno proteine. Na temelju identificiranih proteina napravljena je nutrigenomička analiza iščitavanjem reverzne translacije, odnosno određeno je uključivanje i isključivanje pojedinih metaboličkih putova s poveznicom na promjene nastale u proteomu, kod uzgoja kvasca u aerobnim i anaerobnim uvjetima te pri uskraćivanju i obogaćivanju prihrane kvasca. Utvrđeno je da se nadeksprimirani proteini stanice kvasca razlikuju ovisno o načinu uzgoja, čime se može zaključiti da se ekspresija gena te sinteza proteina, a time i aktivnost metaboličkih putova znatno mijenja pod stresnim promjenama prihrane stanica kvasca.As a result of this thesis, expert system for the management and analysis of MALDI-TOF/TOF generated spectra is developed. This bioinformatics tool serves as a support for the derivatisation and analysis of proteoms of any organism by the use of mass spectrometry. The expert system consists of the proprietary database containing gene and protein data from all publicly available resources with their related taxonomic divisions and sub-divisions. Other inovative part of this system is a graphical user interface (client) used for both input and management of data generated from the spectra analyses. All spectra analyses are carried out on the remote server using algortihms and solutions developed during the course of work on this thesis. As a biological model for testing efficiency for the derivatisation and proteome analysis, the proteome of Saccharomyces cerevisiae was used. S. cerevisiae cells were grown in both aerobic and anaerobic conditions with respective food deprivation as well as optimal food content. Finally, proteins were identified which are with their over or under expression responsible for metabolic changes in yeast under the growth in different environmental conditions