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Content archived on 2024-06-10

Specific labeling of ribosomal particles by dense clusters aimed at phasing crystallographic data

Objective



To understand the molecular basis of protein biosynthesis,
crystallographic studies are being pursued on ribosomes from halophilic and thermophilic bacteria. For phasing by isomorphous replacement methods, dense heavy atoms clusters are being used. An attached undecagold cluster was already shown to possess suitable phasing power, paving the way for the use of a tetrairidium cluster, which by binding to 4-6 sites may lead to phase information at higher resolution.
I plan to specifically derivatize the crystals of the large ribosomal subunits of Haloarcula marismortui (diffracting to 2.9 A), by binding to them, in a few locations, dense and compact clusters. As there are no naturally exposed sulfhydryls on the surface of this particle, I will insert them by site directed mutagenesis after mapping these ribosomal particles and cloning, overproducing and isolating the chosen mutated proteins. Once an inserted sulfhydryl is found to be suitable for binding the clusters, the modified proteins will be incorporated into core particles. I plan to attempt the crystallization of the fully derivatized particles, and if possible to participate in their crystallographic analysis.

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Coordinator

MAX-PLANCK-GESELLSCHAFT ZUR FOERDERUNG DER WISSENSCHAFTEN E.V.
EU contribution
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Address
Ihnestrasse 73
14195 BERLIN
Germany

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Participants (1)

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