α-Conotoxin Decontamination Protocol Evaluation: What Works and What Doesn’t

Nine publically available biosafety protocols for safely handling conotoxin peptides were tested to evaluate their decontamination efficacy. Circular dichroism (CD) spectroscopy and mass spectrometry (MS) were used to assess the effect of each chemical treatment on the secondary and primary structure of α-CTx MII (L10V, E11A). Of the nine decontamination methods tested, treatment with 1% (m/v) solution of the enzymatic detergent Contrex™ EZ resulted in a 76.8% decrease in α-helical content as assessed by the mean residue ellipticity at 222 nm, and partial peptide digestion was demonstrated using high performance liquid chromatography mass spectrometry (HPLC-MS). Additionally, treatment with 6% sodium hypochlorite (m/v) resulted in 80.5% decrease in α-helical content and complete digestion of the peptide. The Contrex™ EZ treatment was repeated with three additional α-conotoxins (α-CTxs), α-CTxs LvIA, ImI and PeIA, which verified the decontamination method was reasonably robust. These results support the use of either 1% Contrex™ EZ solution or 6% sodium hypochlorite in biosafety protocols for the decontamination of α-CTxs in research laboratories

Three-Dimensional Structure of Conotoxin tx3a: A m-1 Branch Peptide of the M-Superfamily

The M-superfamily, one of eight major conotoxin superfamilies found in the venom of the cone snail, contains a Cys framework with disulfide-linked loops labeled 1, 2, and 3 (- CC1C2C3CC-). M-superfamily conotoxins can be divided into the m-1, -2, -3 and -4 branches, based upon the number of residues located in the third Cys loop between the fourth and fifth Cys residues. Here we provide a three-dimensional solution structure for the m-1 conotoxin tx3a found in the venom of Conus textile. The 15 amino acid peptide, CCSWDVCDHPSCTCC, has disulfide bonds between Cys1 and Cys14, Cys2 and Cys12, and Cys7 and Cys15 typical of the C1- C5, C2-C4, and C3-C6 connectivity pattern seen in m-1 branch peptides. The tertiary structure of tx3a was determined by 2D 1H NMR in combination with the combined assignment and dynamics algorithm for nuclear magnetic resonance (NMR) applications CYANA program. Input for structure calculations consisted of 62 inter- and intraproton, 5 phi angle, and 4 hydrogen bond constraints. The root-mean-square deviation values for the 20 final structures are 0.32 +/- 0.07 Å and 0.84 +/- 0.11 Å for the backbone and heavy atoms, respectively. Surprisingly, the structure of tx3a has a “triple-turn” motif seen in the m-2 branch conotoxin mr3a, which is absent in mr3e, the only other member of the m-1 branch of the M-superfamily whose structure is known. Interestingly, injection of tx3a into mice elicits an excitatory response similar to that of the m-2 branch peptide mr3a, even though the conotoxins have different disulfide connectivity patterns

Acceptance Test for Jetstream Test Cluster — Jetstream-Arizona (JA) Dell PowerEdge Test and Development Cluster

This paper details the system description and the performance targets, methods used to perform the acceptance tests, and the achieved performance of the Jetstream test cluster.National Science Foundation Award ACI-1445604: Jetstream - A Self-Provisioned, Scalable Science and Engineering Cloud Environment, Craig A. Stewart, IU, Principal Investigator