Characterization and the biological action mechanism of spider toxin

Research Project

Project/Area Number	60571068
Research Category	Grant-in-Aid for General Scientific Research (C)
Allocation Type	Single-year Grants
Research Field	Biological pharmacy
Research Institution	SETSUNAN UNIVERSITY
Principal Investigator	HOU HIDEMITSU FACULTY OF Pharmaceutical Science, Setsunan University, 薬学部, 助教授 (80101294)
Project Period (FY)	1985 – 1986
Project Status	Completed (Fiscal Year 1986)
Budget Amount *help	¥1,700,000 (Direct Cost: ¥1,700,000) Fiscal Year 1986: ¥400,000 (Direct Cost: ¥400,000) Fiscal Year 1985: ¥1,300,000 (Direct Cost: ¥1,300,000)
Keywords	Joro spider toxin / Purification of JSTX / Glutamate receptor / Glutamate binding / 2,4-Dihydroxyphenylacetic acid
Research Abstract	JSTX which specifically blocked the excitatory postsynaptic potential and glutamate induced potential in crustacean neuromuscular synapses was purified about 300-fold from the venom glands of Nephila clavata by means of column chromatography on Sephadex G-10, CM-Sephadex C-25 and ODS HPLC. Characterization of the purified JSTX we found that the molecular weight was 566 and 2,4-dihydroxyphenylacetic acid, aspartic acid, cadaverine and one unknown component (may be a derivative of lysine) were detected in the acid-hydrolysate. The structure of JSTX was deduced to be Lysine-cadaverine-aspartic acid (asparagine)-2,4-dihydroxyphenylacetic acid. JSTX was shown to inhibit L-glutamate binding to rat brain synaptic membranes in a dose-dependent manner. 2,4-Dihydroxyphenylacetic acid, a common moiety of spider toxins also inhibited specifically L-glutamate binging at a concentration similar to that of the toxin. In addition, the binding activity inhibited by 2,4-dihydroxyphenylacetic acid or JSTX was recovered by the addition of ferric ion. These results suggest that 2,4-dihydroxyphenylacetic acid is a functional part in the molecule of the toxins and the biological action of spider toxin is explained by the direct interaction to Fe-S center which is known to play an important role for the glutamate binding.