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Identification, chemical synthesis, structure, and function of a new K V 1 channel blocking peptide from Oulactis sp.
Author(s) -
Sunanda Punnepalli,
Krishnarjuna Bankala,
Peigneur Steve,
Mitchell Michela L.,
Estrada Rosendo,
VillegasMoreno Jessica,
Pennington Michael W.,
Tytgat Jan,
Norton Raymond S.
Publication year - 2018
Publication title -
peptide science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.533
H-Index - 7
ISSN - 2475-8817
DOI - 10.1002/pep2.24073
Subject(s) - racemization , peptide , cysteine , chemistry , nuclear magnetic resonance spectroscopy , residue (chemistry) , cyclic peptide , peptide sequence , stereochemistry , biochemistry , enzyme , gene
Rapid progress in transcriptomic and proteomic studies of sea anemones has led to the identification of a large number of new peptide sequences. Some of these peptides have high sequence similarity and identical cysteine frameworks to those of previously reported sequences. One such peptide we have identified from a transcriptomic study of Oulactis sp is OspTx2a, which has a cysteine framework similar to that of ShK (from Stichodactyla helianthus ) and BgK (from Bunodosoma granulifera ). This peptide was made using solid‐phase peptide synthesis, but, upon oxidative folding, it generated two peptides with identical masses (OspTx2a‐p1 and OspTx2a‐p2) that were distinguishable by high‐performance liquid chromatography. The structures of OspTx2a‐p1 and OspTx2a‐p2 were determined using nuclear magnetic resonance spectroscopy, and voltage‐clamp electrophysiology assays were performed in order to assess the activity against a range of potassium channels. The structures of the two peptides were very similar to each other and to BgK, with the same disulfide bond connectivities, and both had an all‐ trans backbone conformation. In functional assays, both OspTx2a‐p1 and OspTx2a‐p2 inhibited K V 1.2 and K V 1.6 channel currents at low µM concentrations, with similar but not identical IC 50 values. Peptides containing a C‐terminal Cys residue are particularly sensitive to racemization at this residue, and the two products obtained for OspTx2a could be a consequence of racemization of the Cys residue at its C‐terminus during synthesis. NMR chemical shift differences between the two products and their structural preferences for a d ‐Cys residue were consistent with this interpretation.