A novel-type luciferin from Siberian luminous earthworm Fridericia heliota : structure elucidation by spectral studies and total synthesis

Author Posting. © The Author(s), 2014. This is the author's version of the work. It is posted here by permission of Wiley-VCH Verlag GmbH & Co for personal use, not for redistribution. The definitive version was published in Angewandte Chemie International Edition 53 (2014): 5566–5568, doi:10.1002/anie.201400529.We report structure elucidation and synthesis of the luciferin from the recently discovered luminous earthworm Fridericia heliota. This luciferin represents a key component of a novel ATP-dependent bioluminescence system. The UV, fluorescence, NMR and HRMS spectral studies were performed on 5 mkg of the isolated substance, and gave four isomeric structures, conforming with spectral data. These isomers were chemically synthesized and one of them was found to produce light in the reaction with a protein extract from Fridericia. The novel luciferin was found to have an unusual deeply modified peptidic nature, implying an unprecedented mechanism of action.We acknowledge support from the Program of the Government of the Russian Federation “Measures to attract leading scientists to Russian educational institutions” (grant no. 11. G34.31.0058), the programs MCB RAS, President of the Russian Federation “Leading science school” (grant 3951.2012.4) and the Russian Foundation for Basic Research (grant 14-03-01015). B.M.S. was supported by a stipend from the Program of the President of the Russian Federation.2015-04-1

Impact of membrane partitioning on the spatial structure of an S-type cobra cytotoxin

<p>Cobra cytotoxins (CTs) belong to the three-fingered protein family. They are classified into S- and P-types, the latter exhibiting higher membrane-perturbing capacity. In this work, we investigated the interaction of CTs with phospholipid bilayers, using coarse-grained (CG) and full-atom (FA) molecular dynamics (MD). The object of this work is a CT of an S-type, cytotoxin I (CT1) from <i>N.oxiana</i> venom. Its spatial structure in aqueous solution and in the micelles of dodecylphosphocholine (DPC) were determined by ¹H-NMR spectroscopy. Then, via CG- and FA MD-computations, we evaluated partitioning of CT1 molecule into palmitoyloleoylphosphatidylcholine (POPC) membrane, using the toxin spatial models, obtained either in aqueous solution, or detergent micelle. The latter model exhibits minimal structural changes upon partitioning into the membrane, while the former deviates from the starting conformation, loosing the tightly bound water molecule in the loop-2. These data show that the structural changes elicited by CT1 molecule upon incorporation into DPC micelle take place likely in the lipid membrane, although the mode of the interaction of this toxin with DPC micelle (with the tips of the all three loops) is different from its mode in POPC membrane (primarily with the tip of the loop-1 and both the tips of the loop-1 and loop-2).</p

Structural and Dynamic “Portraits” of Recombinant and Native Cytotoxin I from Naja oxiana: How Close Are They?

Today, recombinant proteins are quite widely used in biomedical and biotechnological applications. At the same time, the question about their full equivalence to the native analogues remains unanswered. To gain additional insight into this problem, intimate atomistic details of a relatively simple protein, small and structurally rigid recombinant cardiotoxin I (CTI) from cobra <i>Naja oxiana</i> venom, were characterized using nuclear magnetic resonance (NMR) spectroscopy and atomistic molecular dynamics (MD) simulations in water. Compared to the natural protein, it contains an additional Met residue at the N-terminus. In this work, the NMR-derived spatial structure of uniformly ¹³C- and ¹⁵N-labeled CTI and its dynamic behavior were investigated and subjected to comparative analysis with the corresponding data for the native toxin. The differences were found in dihedral angles of only a single residue, adjacent to the N-terminal methionine. Microsecond-long MD traces of the toxins reveal an increased flexibility in the residues spatially close to the N-Met. As the detected structural and dynamic changes of the two CTI models do not result in substantial differences in their cytotoxicities, we assume that the recombinant protein can be used for many purposes as a reasonable surrogate of the native one. In addition, we discuss general features of the spatial organization of cytotoxins, implied by the results of the current combined NMR and MD study