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Secondary metabolism and its regulation in Streptomyces ambofaciens: the study of cryptic secondary metabolite biosynthetic gene clusters
Nezbedová, Šárka ; Weiser, Jaroslav (advisor) ; Petříček, Miroslav (referee) ; Mazodier, Philippe (referee)
I. ABSTRACT in English The presented work is focused on secondary metabolism and its regulation in Streptomyces ambofaciens and Streptomyces lividans with special interest in new biosynthetic pathways. The sequencing of bacterial and fungal genomes revealed that the number of their secondary metabolite biosynthetic gene clusters greatly exceeds the number of produced secondary metabolites. Further studies showed that at least some of the newly discovered clusters, called cryptic since no product had been associated to them, were expressed in certain conditions and that they directed the biosynthesis of exploitable secondary metabolites (Gottelt et al., 2010; Gross et al., 2007; Pang et al., 2004). Therefore, these cryptic clusters have been considered as one of the promising reservoirs of new bioactive molecules. Using different approaches I studied the cryptic secondary metabolite biosynthetic gene clusters of Streptomyces ambofaciens, a strain exploited industrially for the production of the antibiotic spiramycin. In the second part of this work, I was interested in the regulation of secondary metabolite biosynthesis and in manipulating regulatory proteins in order to activate the expression of cryptic clusters. The third part of this work studied the effect of the inactivation of the ppk gene, encoding...
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Bacterial biofilm formation
Sochorová, Zuzana ; Zikánová, Blanka (referee) ; Weiser, Jaroslav (advisor)
Biofilms are formed by microorganisms living together in a hydrated extracellular matrix. Formation of such clusters of bacteria brings many benefits. The increased resistance to antibiotics is the main one. Creating a biofilm is analogous to the development of multicellular organisms. Biofilm cells communicate with each other with signaling molecules. Signaling molecules make the biofilm more compact structure. Extracellular matrix, in which bacteria live, maintains biofilm structure, affects cell adhesion and protects cells against environmental influences. Bacteria also interact with the environment through the extracellular matrix. The matrix is composed of various biopolymers and proteins. Biofilms are a common cause of infections associated with implants. There are several ways to prevent bacteria in biofilm to antibiotics. These include a slow diffusion of substances, a slow growth or an adaptation to stress. The formation of persistent cells that are tolerant to the antibiotics is the cause of their survival as well as a new population may arise from them. The surface proteins are important elements for the formation of biofilms, they facilitate adhesion and subsequent establishment of biofilm. A protein Bap was the first identified protein. Later its homologues were found, such as BapA,...
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The study of the biosynthetic pathway of the antibiotic lincomycin
Novotná, Jitka ; Spížek, Jaroslav (advisor) ; Weiser, Jaroslav (referee) ; Gašparík, Juraj (referee)
1. L.3'4-Dihydroxyphenyl a|anine.extraďo|cleavage cyclizationin lincomycin biosynthesis. is followed by intra-molecular The aim of the work gene,characterizeit better DOPA aromaticring is the lincomycinsynthesis. was to assignfunctionto the proteincodedfor by an lmbBl and confirm the assumptionthat2,3-extradiolfission of the actualreactioninvolvedin the metabolicpathwayleadingto fooH HrN-) aOH tyÍo8|n cooH cooH ,,"1 ,,"4 .Ť:+*.i'} 2'3.6ÓcoDoPA /t{3€Íboxy+ox}prcponý}2'34|hyÚ} 1/í pyÍrolc2€íboxy|lo rc|d Fig' 1 |nitia|steps of the amino acid subpathway oÍthe |incomycin biosynthesis The resultsof the feedingexperimentswith labeledintermediatesand subsequent NMR analysis(BRAHME etal., 1984),bearedwitnessof thefactthattheaminoacid sub- pathwayof thelíncomycinbiosynthesisincludes2,3-extradiolcleavageof DoPA (Fig. 1). Neusser and coworkers (NEUSSER et al., 1998) showed that LmbBl catalyzes conversionof DOPA to anunspecifiedyellowcompound. It appearedinapplicableto isolatethe LmbBl reactionproductdirectlyfrom in y,itroreacÍioncatalyzedby thepurifiedLmbBl partlyas LmbBl lost mostof its activity duringdialysis.mostprobablydue to oxidationof theferrousion proposedas a cofactor, andpartlydue to thefact thatmanydiÍ.ferentDoPA oxidationproductswereproducedin the system.Instead,a system simrlar to that applied Íbr...
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