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.
引用
收藏
页码:101 / 108
页数:8
相关论文
共 31 条
  • [21] MCCARTHY WJ, 1987, IN VITRO CELL DEV, V24, P59
  • [22] Means G.E., 1971, CHEM MODIFICATION PR
  • [23] USE OF AZO-DYE-BOUND COLLAGEN TO MEASURE REACTION VELOCITIES OF PROTEOLYTIC ENZYMES
    MOORE, GL
    [J]. ANALYTICAL BIOCHEMISTRY, 1969, 32 (01) : 122 - &
  • [24] STABILITY OF THE LARVICIDAL ACTIVITY OF BACILLUS-THURINGIENSIS SUBSP ISRAELENSIS - AMINO-ACID MODIFICATION AND DENATURANTS
    PFANNENSTIEL, MA
    COUCHE, GA
    MUTHUKUMAR, G
    NICKERSON, KW
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 1985, 50 (05) : 1196 - 1199
  • [25] Riordan J F, 1972, Methods Enzymol, V25, P515, DOI 10.1016/S0076-6879(72)25048-0
  • [26] Riordan J F, 1972, Methods Enzymol, V25, P449, DOI 10.1016/S0076-6879(72)25040-6
  • [27] NUCLEOTIDE-SEQUENCE AND ANALYSIS OF THE N-TERMINAL CODING REGION OF THE SPODOPTERA-ACTIVE DELTA-ENDOTOXIN GENE OF BACILLUS-THURINGIENSIS AIZAWAI-7.29
    SANCHIS, V
    LERECLUS, D
    MENOU, G
    CHAUFAUX, J
    GUO, S
    LECADET, MM
    [J]. MOLECULAR MICROBIOLOGY, 1989, 3 (02) : 229 - 238
  • [28] SCHNEPF HE, 1985, J BIOL CHEM, V260, P6264
  • [29] SHIBANO Y, 1985, GENE, V34, P243
  • [30] PURIFICATION AND CHARACTERIZATION OF THE ACTIVE FRAGMENT FROM BACILLUS-THURINGIENSIS DELTA-TOXIN
    TYSKI, S
    FUJII, Y
    LAI, CY
    [J]. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 1986, 141 (01) : 106 - 111