Dissertation/Thesis Abstract

Studien zu der thaumarchaealen Prolyl-Isomerase NmPin aus Nitrosopumilus maritimus
by Hoppstock, Lukas, Ph.D., Universitaet Duisburg-Essen (Germany), 2016, 110; 10728356
Abstract (Summary)

Peptidyl-prolyl-bonds (Xaa-Pro) are exceptional in peptides and proteins due to their ability to form stable cis-isomers in contrast to all other non-proline peptide bonds. Relative to their position in a protein cis-isomeric Xaa-Pro bonds have a strong impact on the three dimensional structure and function of the protein. Enzymes which catalyse the rotation between cis- and trans-isomers are called peptidyl-prolyl-cis/trans-isomerases (PPIases) and are ubiquitously found in all domains of life. In the cell PPIases play a crucial role in protein folding and protein regulating processes.

This study is the first description of an endogenously expressed and in detail characterized archaeal Parvulin. NmPin from the marine, chemolithoautotrophic thaumarchaeon Nitrosopumilus maritimus is a small parvulin-type single domain PPIase (sdPar) which showed isomerase activity against a subset of unphosphorylated peptidic substrates in a protease-coupled activity assay. By using NMR titration experiments and mutagenesis studies the active site as well as single residues essential for catalysis (D42, S82) were identified in the three dimensional structure of NmPin. In addition a more detailed investigation of the ligand binding mechanism showed the participation of a network of surface exposed, hydrophobic, branched residue sidechains in ligand fixation inside the proline binding pocket.

A thorough topological analysis revealed a positively charged, lysine-rich patch on the protein surface which targets NmPin to lipid vesicles in vitro. The exchange of a single central lysine (K7), which is conserved in thaumarchaeotes, against a glutamate already leads to a loss of the binding ability. In cooperation with the University of Aberdeen immunofluorescence- and transmission electron microscopy of NmPin in N. maritimus cells was done, that confirms that the protein is attached to the outer cell membrane in vivo. These results point towards a PPIase located in a pseudo-perisplamatic space between the cell membrane and the S-Layer. Hence potential cellular substrates for NmPin are proteins secreted into this space or proteins of the S-Layer.

Indexing (document details)
Advisor: Bayer , Peter , Ehrmann, Michael
Commitee:
School: Universitaet Duisburg-Essen (Germany)
School Location: Germany
Source: DAI-C 81/1(E), Dissertation Abstracts International
Source Type: DISSERTATION
Subjects: Microbiology, Biological oceanography
Keywords: Nitrosopumilus maritimus
Publication Number: 10728356
ISBN: 9781392524602
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