The cone snails (genus Conus) are venomous marine molluscs that use small, structured peptide toxins (conotoxins) for prey capture, defense and competitor deterrence. Most biologically active components of these venoms are small peptides, generally 7-35 amino acids in length. Conus peptides generated from genus Conus are organized into families and superfamilies with shared sequence elements. The Conus peptide precursor has a typical signal sequence (the 'pre-region') and a variable spacer (the 'pro-region). Particularly noteworthy is the extreme conservation of signal sequence within all members of a Conus peptide superfamily. The diversity of peptides in different Conus species is apparently generated by hypermutation of the C-terminal mature peptide region. Post-translational modification of Conus peptides requires both specialized enzyme machinery as well as some recognition signal sequence (with a characteristic structural features) in the peptide precursor to instruct the enzymatic machinery as to which amino acid modify. (Image 01)We have observed a D-amino-acid-containing excitatory conotoxin belonging to the I-superfamily that have a D-Leu in the third amino acid from the C-terminus. Post-translational isomerization of L-amino acids to D-amino acids is a subtle modification, not detectable by standard techniques such as Edman sequencing or MS. Accurate predictions require more sequences of modified polypeptides. A noteworthy feature of the I2 superfamily is that it lacks a pro-peptide region between signal sequence and mature toxin, differing from all other conotoxin superfamilies in this respect. The great diversity of I-conopeptide sequences provides a rare opportunity for defining parameters that may be important for this most stealthy of all post-translational modifications. Previous work on post-translational epimerization has indicated that there may be a signal sequence that serves as an anchor site for the modification enzyme. In order to verify if the steric effect of signal sequence serves also to create a hydrophobic reaction environment we have studied the conformational changes of the signal sequence identified for conomarphin with synthetic analogues. Here we report about synthesis and conformational analysis of Boc-VIVFL-OMe and Boc-LLLVIVFL-OMe. Furthermore, a comparison was made between structural properties of Boc-LLLVIVFL-OMe and a D,L-alternating octapeptide containing leucine and valine.
Experimental/Structural study of Conformational Changes in Metabolically Stable Sequences
Emma Fenude
2019
Abstract
The cone snails (genus Conus) are venomous marine molluscs that use small, structured peptide toxins (conotoxins) for prey capture, defense and competitor deterrence. Most biologically active components of these venoms are small peptides, generally 7-35 amino acids in length. Conus peptides generated from genus Conus are organized into families and superfamilies with shared sequence elements. The Conus peptide precursor has a typical signal sequence (the 'pre-region') and a variable spacer (the 'pro-region). Particularly noteworthy is the extreme conservation of signal sequence within all members of a Conus peptide superfamily. The diversity of peptides in different Conus species is apparently generated by hypermutation of the C-terminal mature peptide region. Post-translational modification of Conus peptides requires both specialized enzyme machinery as well as some recognition signal sequence (with a characteristic structural features) in the peptide precursor to instruct the enzymatic machinery as to which amino acid modify. (Image 01)We have observed a D-amino-acid-containing excitatory conotoxin belonging to the I-superfamily that have a D-Leu in the third amino acid from the C-terminus. Post-translational isomerization of L-amino acids to D-amino acids is a subtle modification, not detectable by standard techniques such as Edman sequencing or MS. Accurate predictions require more sequences of modified polypeptides. A noteworthy feature of the I2 superfamily is that it lacks a pro-peptide region between signal sequence and mature toxin, differing from all other conotoxin superfamilies in this respect. The great diversity of I-conopeptide sequences provides a rare opportunity for defining parameters that may be important for this most stealthy of all post-translational modifications. Previous work on post-translational epimerization has indicated that there may be a signal sequence that serves as an anchor site for the modification enzyme. In order to verify if the steric effect of signal sequence serves also to create a hydrophobic reaction environment we have studied the conformational changes of the signal sequence identified for conomarphin with synthetic analogues. Here we report about synthesis and conformational analysis of Boc-VIVFL-OMe and Boc-LLLVIVFL-OMe. Furthermore, a comparison was made between structural properties of Boc-LLLVIVFL-OMe and a D,L-alternating octapeptide containing leucine and valine.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
