Purification of an inositol 1,4,5-trisphosphate-binding calreticulin-containing intracellular compartment of HL-60 cells

C. Van Delden, C. Favre, A. Spät, E. Cerny, K. H. Krause, D. P. Lew

Research output: Contribution to journalArticle

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Abstract

To investigate the identity of Ins(1,4,5)P3-sensitive intracellular Ca2+ stores in myeloid cells, we have developed a method that yields subcellular fractions highly enriched in Ins(1,4,5)P3 binding. HL-60 cells were disrupted by nitrogen cavitation, and subcellular fractions were obtained by differential centrifugation, followed by Percoll- and sucrose-density-gradient separations. A subcellular fraction enriched 26-fold in Ins(1,4,5)P3-binding sites was obtained. This fraction showed no enrichment in plasma-membrane markers and only a comparatively moderate enrichment (7-fold) in endoplasmic-reticulum markers. The ratio between specific enrichment of Ins(1,4,5)P3 binding and endoplasmic-reticulum markers in the different fractions varied over 50-fold, from <0.1 to > 5. The purified Ins(1,4,5)P3-binding fraction was enriched to a similar extent (27-fold) in the putative intravesicular Ca2+ -storage protein calreticulin. Our results favour the concept of a distinct Ins(1,4,5)P3-binding, calreticulin-containing compartment (i.e. the calciosome) in HL-60 cells.

Original languageEnglish
Pages (from-to)651-656
Number of pages6
JournalBiochemical Journal
Volume281
Issue number3
Publication statusPublished - 1992

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Calreticulin
Inositol 1,4,5-Trisphosphate
Subcellular Fractions
HL-60 Cells
Purification
Endoplasmic Reticulum
Centrifugation
Cell membranes
Cavitation
Sucrose
Nitrogen
Binding Sites
Myeloid Cells
Cell Membrane
Proteins
Percoll

ASJC Scopus subject areas

  • Biochemistry

Cite this

Purification of an inositol 1,4,5-trisphosphate-binding calreticulin-containing intracellular compartment of HL-60 cells. / Van Delden, C.; Favre, C.; Spät, A.; Cerny, E.; Krause, K. H.; Lew, D. P.

In: Biochemical Journal, Vol. 281, No. 3, 1992, p. 651-656.

Research output: Contribution to journalArticle

Van Delden, C. ; Favre, C. ; Spät, A. ; Cerny, E. ; Krause, K. H. ; Lew, D. P. / Purification of an inositol 1,4,5-trisphosphate-binding calreticulin-containing intracellular compartment of HL-60 cells. In: Biochemical Journal. 1992 ; Vol. 281, No. 3. pp. 651-656.
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AU - Krause, K. H.

AU - Lew, D. P.

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