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Light-harvesting like domain of the cyanobacterial ferrochelatase
PAZDERNÍK, Marek
This thesis is focused on elucidating the function of the C-terminal transmembrane lightharvesting complex like (LHC) domain of the cyanobacterial ferrochelatase (FeCh). Using the model cyanobacterium Synechocystis PCC 6803, I show that the FeCh LHC domain can bind chlorophyll (Chl) and carotenoids; however, this pigment binding occurs only when the biosynthesis of heme and Chl in the cell is misbalanced. Further, I found that point mutation, which prevents the pigment binding to FeCh LHC domain results in a misregulated ratio between heme and Chl during stress conditions due to low heme accumulation. My data also show that the FeCh LHC domain interacts with CurT protein most likely to localize the FeCh into a specialized membrane domain, where the synthesis of photosystem II is proposed to occur. Based on my data I propose that the role of the FeCh LHC domain is to monitor the availability of Chl during photosystem biogenesis and to coordinate Chl availability with the synthesis of heme.
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Molecular pathology of selected porphyria with skin manifestation
Sameh Anwar Hussein Farrag, Mohamed ; Martásek, Pavel (advisor) ; Baxová, Alice (referee) ; Raman, C. S. (referee)
Porphyria is a group of inherited metabolic disorders due to enzymatic defect of the heme biosynthesis resulting in the overproduction of the heme precursors' porphyrins in different body organs. The enzymes of the heme biosynthesis are encoded by corresponding genes in which any defect in any of these genes lead to a specific type of porphyria. Numerous mutations were detected in these genes leading to impairment in the enzyme function and therefore developing of the clinical manifestations of porphyria. The aim of the present work was to investigate the UROD gene in patients with porphyria cutanea tarda (PCT) and hepatoerythropoietic protoporphyria (HEP) as well as the FECH gene in patients with erythropoietic protoporphyria (EPP) on a molecular level. We identified numerous mutations in the FECH and the UROD genes in three different populations, Czech, Slovak, and Egyptian. We described the novel mutations in the UROD gene in HEP Arabic patients from Egypt as well in the FECH gene in patients with EPP of Czech and Slovak origin. We expressed mutatted UROD protein in prokaryotic system and found 19 % of the wild-type enzymatic activity. Moreover, the current study presents for the first time the frequency of the low expression allele IVS3-48c in the FECH gene in healthy controls from the Czech...
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Molecular pathology of selected porphyria with skin manifestation
Sameh Anwar Hussein Farrag, Mohamed ; Martásek, Pavel (advisor) ; Baxová, Alice (referee) ; Raman, C. S. (referee)
Porphyria is a group of inherited metabolic disorders due to enzymatic defect of the heme biosynthesis resulting in the overproduction of the heme precursors' porphyrins in different body organs. The enzymes of the heme biosynthesis are encoded by corresponding genes in which any defect in any of these genes lead to a specific type of porphyria. Numerous mutations were detected in these genes leading to impairment in the enzyme function and therefore developing of the clinical manifestations of porphyria. The aim of the present work was to investigate the UROD gene in patients with porphyria cutanea tarda (PCT) and hepatoerythropoietic protoporphyria (HEP) as well as the FECH gene in patients with erythropoietic protoporphyria (EPP) on a molecular level. We identified numerous mutations in the FECH and the UROD genes in three different populations, Czech, Slovak, and Egyptian. We described the novel mutations in the UROD gene in HEP Arabic patients from Egypt as well in the FECH gene in patients with EPP of Czech and Slovak origin. We expressed mutatted UROD protein in prokaryotic system and found 19 % of the wild-type enzymatic activity. Moreover, the current study presents for the first time the frequency of the low expression allele IVS3-48c in the FECH gene in healthy controls from the Czech...
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Expression and purification of \kur{Synechocystis} ferrochelatase from \kur{Escherichia coli}
RICHTOVÁ, Jitka
Ferrochelatase (FeCH) is an ubiquitous enzyme producing heme, an essential pigment for all forms of organisms. In photosynthetic organism, heme is synthesized together with the chlorophyll in one branched pathway and the FeCH enzyme appears to be important for regulation of both the chlorophyll and the heme biosynthesis. To understand regulatory role of this protein, an active recombinant FeCH from photosynthetic organism would be invaluable. The aim of this project is to express FeCH from cyanobacterium Synechocystis 6803 in Escherichia coli and to prepare a protocol for the purification of this protein as a highly active enzyme.
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Evolution of selected enzymes of the shikimate pathway and the haem biosynthetic pathway in Rhodophyta (class Florideophyceae)
VORÁČOVÁ, Kateřina
Diatoms derived their plastid from red algae through the secondary endosymbiosis. Most of the endosymbiont genes have been transferred from the engulfed alga to the secondary host nucleus, therefore evolution of these genes correspond to the evolution of plastids rather than to the evolution of the host organisms. Similarly, genes coding for ferrochelatase and DAHP synthase from diatoms are closely related to those from plants and green algae. Contrary to this, red algal genes do not cluster within this clade. I tried to amplify and sequence genes coding for ferrochelatase and DAHP synthase from representatives of the class Florideophyceae to investigate their phylogenetic position.
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