No description available.

Universiẗat, Diss., 2001--Freiburg (Breisgau).

No description available.

<p>5’-Nucleotidases (5’NTs) are catabolic enzymes of the nucleotide metabolism. They catalyze dephosphorylation of deoxyribo- and ribonucleoside monophosphates and constitute an important control point in the regulation of intracellular nucleotide pools for the maintenance of correct DNA and RNA synthesis.</p><p>By removing the alfa-phosphate group from a nucleotide, the 5’NTs release the nucleoside to pass the plasma membrane by facilitated diffusion. Depending on the cellular need for nucleotides, the nucleosides can either exit the cell for reuse elsewhere or be imported and subsequently phosphorylated by nucleoside and nucleotide kinases.</p><p>The knowledge of how nucleotides are metabolized has been used for rational design of nucleoside analogues that are used in treatment of cancer and viral diseases. These drugs are phosphorylated within the cell to become active. Their dephosphorylation by 5’NTs might be one of the mechanisms behind the resistance experienced by patients towards such drugs.</p><p>This thesis describes structure-function studies on four of the seven known human 5’-NTs. The focus of the work is on the substrate specificity and regulation of these enzymes. Inactive variants of the mitochondrial and cytosolic deoxynucleotidases and the cytosolic 5’-nucleotidase II were used to characterize the structural basis for their substrate specificity in high detail.</p><p>Based on structures of the apoprotein and activator/activator+substrate complexes of cytosolic 5’-nucleotidase II, a mechanism for the allosteric activation of this enzyme was presented. In this mechanism, the activator induces a conformational change that involves conserved residues of the active site. The conformational change drastically increases the enzyme affinity for the phosphate moiety of the substrate.</p>

5’-Nucleotidases (5’NTs) are catabolic enzymes of the nucleotide metabolism. They catalyze dephosphorylation of deoxyribo- and ribonucleoside monophosphates and constitute an important control point in the regulation of intracellular nucleotide pools for the maintenance of correct DNA and RNA synthesis. By removing the alfa-phosphate group from a nucleotide, the 5’NTs release the nucleoside to pass the plasma membrane by facilitated diffusion. Depending on the cellular need for nucleotides, the nucleosides can either exit the cell for reuse elsewhere or be imported and subsequently phosphorylated by nucleoside and nucleotide kinases. The knowledge of how nucleotides are metabolized has been used for rational design of nucleoside analogues that are used in treatment of cancer and viral diseases. These drugs are phosphorylated within the cell to become active. Their dephosphorylation by 5’NTs might be one of the mechanisms behind the resistance experienced by patients towards such drugs. This thesis describes structure-function studies on four of the seven known human 5’-NTs. The focus of the work is on the substrate specificity and regulation of these enzymes. Inactive variants of the mitochondrial and cytosolic deoxynucleotidases and the cytosolic 5’-nucleotidase II were used to characterize the structural basis for their substrate specificity in high detail. Based on structures of the apoprotein and activator/activator+substrate complexes of cytosolic 5’-nucleotidase II, a mechanism for the allosteric activation of this enzyme was presented. In this mechanism, the activator induces a conformational change that involves conserved residues of the active site. The conformational change drastically increases the enzyme affinity for the phosphate moiety of the substrate.

In duplex DNA, guanine radical cations react with water to form mainly 7,8-dihydro-8-oxoguanine (8-OxoG). Understanding for the mechanism for migration of a radical cation (hole) from the site of initial DNA oxidation to a remote guanine is an important step in the process that will lead to a thorough understanding of DNA damage and its repair. The vast majority of charge migration in DNA experiments utilize guanine oxidation as a monitor for charge transfer. The synthesis of a potential monitor for charge transfer through DNA that is independent of guanine oxidation is reported herein. The system is a carbazole moiety covalently attached to the 2O position of uridine which was successfully incorporated into a DNA strand. Carbazole has a low oxidation potential, and will create a deeper trap than guanine during DNA charge transfer. One electron oxidation of carbazole should lead to the formation of its radical cation. The high extinction coefficient of carbazole radical cation should make it clearly observable with UV analysis. Hence a monitor for charge migration in DNA independent of guanine oxidation is obtained.

No / The genome of Caenorhabditis elegans encodes multiple homologues of the two major families of mammalian equilibrative and concentrative nucleoside transporters. As part of a programme aimed at understanding the biological rationale underlying the multiplicity of eukaryote nucleoside transporters, we have now demonstrated that the nematode genes ZK809.4 (ent-1) and K09A9.3 (ent-2) encode equilibrative transporters, which we designate CeENT1 and CeENT2 respectively. These transporters resemble their human counterparts hENT1 and hENT2 in exhibiting similar broad permeant specificities for nucleosides, while differing in their permeant selectivities for nucleobases. They are insensitive to the classic inhibitors of mammalian nucleoside transport, nitrobenzylthioinosine, dilazep and draflazine, but are inhibited by the vasoactive drug dipyridamole. Use of green fluorescent protein reporter constructs indicated that the transporters are present in a limited number of locations in the adult, including intestine and pharynx. Their potential roles in these tissues were explored by using RNA interference to disrupt gene expression. Although disruption of ent-1 or ent-2 expression alone had no effect, simultaneous disruption of both genes yielded pronounced developmental defects involving the intestine and vulva.

No description available.

Thesis (doctoral)--Universität, Tübingen, 2003.

No description available.