Parabutoporin (PP) and opistoporin 1 (OP1) are cation, a-helical antimicrobial peptides isolated
from the southern African scorpion species, Parabuthus schlechteri and Opistophthalmus carinatus,
respectively. Along with their antimicrobial action against bacteria and fungi, these peptides show
pore-forming properties in the membranes of mammalian cells. Pore-formation and ion selectivity in
cardiac myocytes were investigated by measuring the whole cell leak current by means of the patch
clamp technique. Pore-formation was observed as the induction of leak currents. Ion selectivity of
the pores was indicated by the shift of the reversal potential (E,,,) upon substitution of intra (K' with
CS' and CI- with aspartate) and extracellular (Na' with NMDG') ions. Results were compared with
the effect of gramicidin A used as a positive control for monovalent cation selective pores. PP and
OP I induced a fluctuating leak current and indicate non-selectivity of PP and OP1-induced pores.
An osmotic protection assay to determine estimated pore size was performed on the cardiac
myocytes. PP and OP1-induced pores had an estimate pore size of 1.38-1.78 nm in diameter.
The effect of PP and OP1 on the membrane potential (MP) of a neuroblastoma cell line and cardiac
myocytes was investigated. TMRM was used to mark the MP fluorescently and a confocal
microscope used to record the data digitally. The resting membrane potential (RMP) of the
neuroblastoma cells was calculated at -38.3 f 1.9 mV. PP (0.5 uM) and OP1 (0.5-1 uM) depolarized
the entire cell uniformly to a MP of -1 1.9 k 3.9 mV and -9.4 k 1.9 mV, respectively. This occurred
after 20-30 min of peptide exposure. In the case of the cardiac myocytes depolarization was induced
to -39.7 f 8.4 mV and -32.6 f 5.2 mV by 0.5-1 uM PP and 1.5-2.5 uM OPl, respectively. / Thesis (M.Sc. (Physiology))--North-West University, Potchefstroom Campus, 2006.
| Identifer | oai:union.ndltd.org:NWUBOLOKA1/oai:dspace.nwu.ac.za:10394/991 |
| Date | January 2005 |
| Creators | Elgar, Dale |
| Publisher | North-West University |
| Source Sets | North-West University |
| Detected Language | English |
| Type | Thesis |
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