Saturday, October 13, 2012: 10:00 PM
Hall 4E/F (WSCC)
Melanoma is the most malignant type of skin cancer. Most melanomas do not respond to
chemotherapies and surgery is the most feasible treatment available. Mojastin, a disintegrin found in
venom of the Mojave rattlesnake, can alter cell signaling by binding to integrins. This is primarily due to
an RGD binding motif, followed by an amino acid sequence that can induce various cellular responses.
In previous research, our research group demonstrated that r-Moj-DM induced apoptosis of SK-Mel-28
cells, a human melanoma cell line, while the wild type r-Moj-WN and r-Moj-NN did not. Our hypothesis
is that altering the tryptophan (W) to an aspartate (D) carboxyl of the RGD is sufficient to induce
apoptosis of Sk-Mel-28 cells. In order to test this hypothesis we produced three mutants (r-Moj-DL, r-
Moj-DN, and r-Moj-DP) using site-directed mutagenesis. The recombinant peptides were expressed and
purified using GST bead purification and thrombin cleavage. SK-Mel-28 cells were cultured on chamber
slides treated with r-Moj-DL, r-Moj-DN, or PBS as a negative control. Cells treated with r-Moj-DL showed
chromatin fragmentation while the results of r-Moj-DN were inconclusive. Nuclear fragmentation
experiments with r-Moj-DP and r-Moj-DN will be performed. In addition, tunnel assays will be used to
quantitate the number of cells undergoing apoptosis after r-Moj-DL, r-Moj-DN, and r-Moj-DP treatment.
Our preliminary data with the r-Moj-DL may support our hypothesis.
chemotherapies and surgery is the most feasible treatment available. Mojastin, a disintegrin found in
venom of the Mojave rattlesnake, can alter cell signaling by binding to integrins. This is primarily due to
an RGD binding motif, followed by an amino acid sequence that can induce various cellular responses.
In previous research, our research group demonstrated that r-Moj-DM induced apoptosis of SK-Mel-28
cells, a human melanoma cell line, while the wild type r-Moj-WN and r-Moj-NN did not. Our hypothesis
is that altering the tryptophan (W) to an aspartate (D) carboxyl of the RGD is sufficient to induce
apoptosis of Sk-Mel-28 cells. In order to test this hypothesis we produced three mutants (r-Moj-DL, r-
Moj-DN, and r-Moj-DP) using site-directed mutagenesis. The recombinant peptides were expressed and
purified using GST bead purification and thrombin cleavage. SK-Mel-28 cells were cultured on chamber
slides treated with r-Moj-DL, r-Moj-DN, or PBS as a negative control. Cells treated with r-Moj-DL showed
chromatin fragmentation while the results of r-Moj-DN were inconclusive. Nuclear fragmentation
experiments with r-Moj-DP and r-Moj-DN will be performed. In addition, tunnel assays will be used to
quantitate the number of cells undergoing apoptosis after r-Moj-DL, r-Moj-DN, and r-Moj-DP treatment.
Our preliminary data with the r-Moj-DL may support our hypothesis.