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Isolation of Translating Ribosomes Containing Peptidyl-tRNAs for Functional and Structural Analyses
Published on: February 26, 2011
ATPase activity of non-ribosomal peptide synthetases
Maja Pavela-Vrancic1, Ralf Dieckmann, Hans von Döhren
1Department of Chemistry, Faculty of Natural Sciences, Mathematics and Education, University of Split, N Tesle 12, 21000 Split, Croatia.
Abstract:
Adenylation domains of non-ribosomal peptide synthetases (NRPS) catalyse the formation of aminoacyl adenylates, and in addition synthesize mono- and dinucleoside polyphosphates. Here, we show that NRPS systems furthermore contain an ATPase activity in the range of up to 2 P(i)/min. The hydrolysis rate by apo-tyrocidine synthetase 1 (apo-TY1) is enhanced in the presence of non-cognate amino acid substrates, correlating well with their structural features and the diminishing adenylation efficiency. A comparative analysis of the functional relevance of an analogous sequence motif in P-type ATPases and adenylate kinases (AK) allowed a putative assignment of the invariant aspartate residue from the TGDLA(V)R(K) core sequence in NRPS as the Mg(2+) binding site. Less pronounced variations in ATPase activity are observed in domains with relaxed amino acid specificity of gramicidin S synthetase 2 (GS2) and delta-(L-aminoadipyl)-L-cysteinyl-D-valine synthetase (ACVS), known to produce a set of substitutional variants of the respective peptide product. These results disclose new perspectives about the mode of substrate selection by NRPS.
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