CHEMICAL MODIFICATION OF BACILLUS-THURINGIENSIS SUBSP THURINGIENSIS (HD-524) TRYPSIN-ACTIVATED ENDOTOXIN - IMPLICATION OF TYROSINE RESIDUES IN LEPIDOPTERAN CELL-LYSIS

被引:10
作者
YAN, XJ [1 ]
MCCARTHY, WJ [1 ]
机构
[1] PENN STATE UNIV,DEPT ENTOMOL,PESTICIDE RES LAB,UNIVERSITY PK,PA 16802
关键词
LEPIDOPTERAN CELL CULTURE; BACILLUS-THURINGIENSIS SUBSP THURINGIENSIS LYSIS OF; BACILLUS-THURINGIENSIS SUBSP THURINGIENSIS TOXIN; AMINO ACID MODIFICATION OF BACILLUS-THURINGIENSIS TOXIN;
D O I
10.1016/0022-2011(91)90046-S
中图分类号
Q95 [动物学];
学科分类号
071002 ;
摘要
A purified protein fraction from a solubilized and trypsin-digested extract of Bacillus thuringiensis subsp. thuringiensis (HD-524) fermentation powder was lytic to cells from several lepidopteran lines. Maximum yield was obtained by alkaline carbonate-thiocyanate solubilization of washed powder followed by trypsin digestion and Sephacryl (S-300) chromatography. The alkaline carbonate-solubilized fraction consisted predominantly of two bands on sodium dodecyl sulfatepolyacrylamide gel electrophoresis with MW of 144 ± 0.9 kDa and 134 ± 1.4 kDa. After trypsin treatment and column chromatography, the cytolytic fraction consisted of a major band with a MW of 60.0 ± 1.8 kDa and a minor band of 69 ± 0.9 kDa. Cells from Trichoplusia ni (TN368) were most susceptible to lysis with 50% of cells lysed at 3 μg/ml, followed by Spodoptera frugiperda cells (SF21AE) exhibiting 50% cell lysis at 5 μg/ml and Lymantria dispar cells (Ld652Y) showing 40% lysis at 10 μg/ml. Chemical modification of the polypeptides was performed to determine the role of certain amino acid residues in the cytolytic activity. The group-specific reagent tetranitromethane was used to nitrate and oxidize tyrosine and cysteine residues, respectively. Cysteine residues alone were also modified with p-hydroxymercuribenzoic acid. Lysine residues were modified with O-methylisourea. Of the three types of amino acid residues, only the modification of tyrosine resulted in reduced cell lysis. © 1991.
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页码:101 / 108
页数:8
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