NMR resonance assignments for a docking domain pair with an attached thiolation domain from the PAX peptide-producing NRPS from Xenorhabdus cabanillasii

Non-ribosomal peptide synthetases (NRPSs) are large multienzyme machineries. They synthesize numerous important natural products starting from amino acids. For peptide synthesis functionally specialized NRPS modules interact in a defined manner. Individual modules are either located on a single or on multiple different polypeptide chains. The “peptide-antimicrobial-Xenorhabdus” (PAX) peptide producing NRPS PaxS from Xenorhabdus bacteria consists of the three proteins PaxA, PaxB and PaxC. Different docking domains (DDs) located at the N-termini of PaxB and PaxC and at the C-termini of PaxA and BaxB mediate specific non-covalent interactions between them. The N-terminal docking domains precede condensation domains while the C-terminal docking domains follow thiolation domains. The binding specificity of individual DDs is important for the correct assembly of multi-protein NRPS systems. In many multi-protein NRPS systems the docking domains are sufficient to mediate the necessary interactions between individual protein chains. However, it remains unclear if this is a general feature for all types of structurally different docking domains or if the neighboring domains in some cases support the function of the docking domains. Here, we report the 1H, 13C and 15 N NMR resonance assignments for a C-terminal di-domain construct containing a thiolation (T) domain followed by a C-terminal docking domain (CDD) from PaxA and for its binding partner – the N-terminal docking domain (NDD) from PaxB from the Gram-negative entomopathogenic bacterium Xenorhabdus cabanillasii JM26 in their free states and for a 1:1 complex formed by the two proteins. These NMR resonance assignments will facilitate further structural and dynamic studies of this protein complex.

NMR resonance assignments of the FinO-domain of the RNA chaperone RocC

In prokaryotic species, gene expression is commonly regulated by small, non-coding RNAs (sRNAs). In the gram-negative bacterium Legionella pneumophila, the regulatory, trans-acting sRNA molecule RocR base pairs with a complementary sequence in the 5’-untranslated region of mRNAs encoding for proteins in the bacterial DNA uptake system, thereby controlling natural competence. Sense-antisense duplexing of RocR with targeted mRNAs is mediated by the recently described RNA chaperone RocC. RocC contains a 12 kDa FinO-domain, which acts as sRNA binding platform, along with an extended C-terminal segment that is predicted to be mostly disordered but appears to be required for repression of bacterial competence. In this work we assigned backbone and side chain 1H, 13C, and 15N chemical shifts of RocC’s FinO-domain by solution NMR spectroscopy. The chemical shift data for this protein indicate a mixed α/β fold that is reminiscent of FinO from Escherichia coli. Our NMR resonance assignments provide the basis for a comprehensive analysis of RocC’s chaperoning mechanism on a structural level.