Höcker, Birte and Claren, Jörg and Sterner, Reinhard (2004) Mimicking enzyme evolution by generating new (betaalpha)8-barrels from (betaalpha)4-half-barrels. Proceedings of the National Academy of Sciences of the United States of America 101 (47), pp. 16448-53.
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Gene duplication and fusion events that multiply and link functional protein domains are crucial mechanisms of enzyme evolution. The analysis of amino acid sequences and three-dimensional structures suggested that the (betaalpha)8-barrel, which is the most frequent fold among enzymes, has evolved by the duplication, fusion, and mixing of (betaalpha)4-half-barrel domains. Here, we mimicked this evolutionary strategy by generating in vitro (betaalpha)8-barrels from (betaalpha)4-half-barrels that were deduced from the enzymes imidazole glycerol phosphate synthase (HisF) and N'[(5'-phosphoribosyl)formimino]-5-aminoimidazole-4-carboxamide-ribonucleotide isomerase (HisA). To this end, the gene for the C-terminal (betaalpha)4-half-barrel (HisF-C) of HisF was duplicated and fused in tandem to yield HisF-CC, which is more stable than HisF-C. In the next step, by optimizing side-chain interactions within the center of the beta-barrel of HisF-CC, the monomeric and compact (betaalpha)8-barrel protein HisF-C*C was generated. Moreover, the genes for the N- and C-terminal (betaalpha)4-half-barrels of HisF and HisA were fused crosswise to yield the chimeric proteins HisFA and HisAF. Whereas HisFA contains native secondary structure elements but adopts ill-defined association states, the (betaalpha)8-barrel HisAF is a stable and compact monomer that reversibly unfolds with high cooperativity. The results obtained suggest a previously undescribed dimension for the diversification of enzymatic activities: new (betaalpha)8-barrels with novel functions might have evolved by the exchange of (betaalpha)4-half-barrel domains with distinct functional properties.
|Institutions:||Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Reinhard Sterner|
|Subjects:||500 Science > 570 Life sciences|
|Refereed:||Yes, this version has been refereed|
|Created at the University of Regensburg:||Yes|
|Deposited On:||22 Mar 2010 08:15|
|Last Modified:||22 Mar 2010 08:15|