Characterization of azadirachtin binding to Sf9 nuclei in vitro

Alasdair J. Nisbet, A. Jennifer Mordue (Luntz), Robert B. Grossman, Lyn Jennens, Steven V. Ley, William Mordue

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

[22,23-(3)H(2)]dihydroazadirachtin was incorporated by Sf9 cells in culture and was bound specifically to the nuclear fraction. The observed association constant of the binding of the radioligand to a purified nuclear fraction was determined to be 0.037 +/- 0.008 min(-1) using a one-phase exponential association equation, and binding appeared to be to a single population of sites. The binding was essentially irreversible, and the dissociation constant was estimated to be 0.00065 +/- 0.00013 min(-1). An association rate constant of 7.3 x 10(6) M(-1) min(-1) was calculated from these data. Binding was saturable, and the receptor number and affinity were determined as B(max) = 23.87 +/- 1.15 pmol/mg protein, K(d) = 18.1 +/- 2.1 nM. The order of potency of semisynthetic azadirachtin analogues for competition for the binding site was as follows (IC(50) in parentheses): azadirachtin (1.55 x 10(-8) M) > dihydroazadirachtin (3.16 x 10(-8) M) > dansyl dihydroazadirachtin (7.40 x 10(-8) M) > DNP-azadirachtin (7.50 x 10(-8) M) > biotin dihydroazadirachtin (1.27 x 10(-7) M) >> 11-methoxy 22,23-dihydroazadirachtin (6.67 x 10(-7) M). [Originally published in Volume 34, Archives of Insect Biochemistry and Physiology, 34:461-473 (1997).]

Original languageEnglish
Pages (from-to)78-86
Number of pages9
JournalArchives of Insect Biochemistry and Physiology
Volume46
Issue number1-2
DOIs
StatePublished - 2001

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Insect Science

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