TY  - THES
T1  - Metal and inhibitor binding studies on metallo-beta-lactamases
A1  - Selevsek,Nathalie
Y1  - 2009/01/26
N2  - The heterogeneity of the metal content observed in Metallo-ß-Lactamases (MBLs) hampers the design of potential inhibitors. In the first part of the work, three representative members of the MBLs, namely BcII, CphA and L1 were investigated using mass spectrometric and spectroscopic methods. Experimental parameters for the detection of the metal-protein and ternary metalloprotein-inhibitor complexes using ESI-MS1 were evaluated and optimized. SAR1 determined in the gas phase were in agreement with kinetic assays performed in solution. This demonstrates the suitability of this technique for the screening for new inhibitors of MBLs and for the detection of metal:enzyme:inhibitor ratios. Competition-titrations in combination with ESI-MS, revealed that for different subclasses of the MBL, the inhibition by (R,S)-thiomandelate and D-captopril is strongly influenced by the nature of the metal ion and the metal content of the protein. In the second part of the work, the metal ion dependent flexibility of different parts of the BcII protein was investigated using HDX-MS1. It was shown that the metal-free enzyme was the least ordered structure and that the high flexibility at the metal binding site and the domain interface region in the Cd1-enzyme might facilitate the transfer of the metal between the two binding sites. These findings deliver important parameters for future development of efficient inhibitors for these enzymes.
KW  - Lactamase <beta->
KW  - Struktur-Aktivitäts-Beziehung
KW  - Massenspektrometrie
KW  - Elektronensprayionisations-Massenspektrometrie
KW  - MALDI-MS
CY  - Saarbrücken
PB  - Universitäts- und Landesbibliothek
AD  - Postfach 151141, 66041 Saarbrücken
UR  - http://scidok.sulb.uni-saarland.de/volltexte/2009/2035
ER  -