Peterhoff, David and Zellner, Hermann and Guldan, Harald and Merkl, Rainer and Sterner, Reinhard and Babinger, Patrick (2012) Dimerization Determines Substrate Specificity of a Bacterial Prenyltransferase. CHEMBIOCHEM, 13 (9). pp. 1297-1303. ISSN 1439-4227, 1439-7633
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We have identified the native dimer interface of heptaprenylglyceryl phosphate synthase PcrB from the bacterium Bacillus subtilis and analyzed the significance of oligomer formation for stability and catalytic activity. Computational methods predicted two different surface regions of the PcrB protomer that could be responsible for dimer formation. These bona fide interfaces were assessed both in silico and experimentally by the introduction of amino acid substitutions that led to monomerization, and by incorporation of an unnatural amino acid to allow cross-linking of the two protomers. The results showed that, in contrast to previous assumptions, PcrB uses the same interface for dimerization as the homologous geranylgeranylglyceryl phosphate synthase from Archaea. Thermal unfolding demonstrated that the monomeric proteins are only slightly less stable than wild-type PcrB. However, activity assays showed that monomerization limits the length of accepted polyprenyl pyrophosphates to three isoprene units, whereas the native PcrB substrate contains seven isoprene entities. We provide a plausible hypothesis as to how dimerization determines substrate specificity of PcrB.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | GERANYLGERANYLGLYCERYL PHOSPHATE SYNTHASE; CHAIN-LENGTH DETERMINATION; SITE-DIRECTED MUTAGENESIS; LINKING AMINO-ACID; ESCHERICHIA-COLI; DIPHOSPHATE SYNTHASE; CRYSTAL-STRUCTURE; GENETIC-CODE; ENZYME; PROTEIN; enzymes; ether lipids; oligomerization; protein engineering; unnatural amino acids |
| Subjects: | 500 Science > 570 Life sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Reinhard Sterner Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Rainer Merkl |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 12 May 2020 05:44 |
| Last Modified: | 12 May 2020 05:44 |
| URI: | https://pred.uni-regensburg.de/id/eprint/18572 |
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