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PMID: 2829827 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Characterization of partially purified phospholipase C from human platelet membranes.

The Biochemical journal ·Vol. 248 ·No. 1 ·1987-11-15 ·Pages 95-101

Banno Y, Nozawa Y

Abstract

A phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2]-hydrolytic activity was found to be present in the human platelet membrane fraction, with 20% of the total activity of the homogenate. The membrane-associated phospholipase C activity was extracted with 1% deoxycholate (DOC). The DOC-extractable phospholipase C was partially purified approx. 126-fold to a specific activity of 0.58 mumol of PtdIns-(4,5)P2 cleaved/min per mg of protein, by Q-Sepharose, heparin-Sepharose and Ultrogel AcA-44 column chromatographies. This purified DOC-extractable phospholipase C had an Mr of approx. 110,000, as determined by Ultrogel AcA-44 gel filtration. The enzyme exhibits a maximal hydrolysis for PtdIns-(4,5)P2 at pH 6.5 in the presence of 0.1% DOC. The addition of 0.1% DOC caused a marked activation of both PtdIns(4,5)P2 and phosphatidylinositol (PtdIns) hydrolyses by the enzyme. The enzyme hydrolysed PtdIns(4,5)P2 and PtdIns in a different Ca2+-dependent manner; the maximal hydrolyses for PtdIns(4,5)P2 and PtdIns were obtained at 4 microM- and 0.5 mM-Ca2+ respectively. In the presence of 1 mM-Mg2+, PtdIns(4,5)P2-hydrolytic activity was decreased at all Ca2+ concentrations examined, but PtdIns-hydrolytic activity was not affected.

MeSH Terms
Blood Platelets/drug effects,enzymology Calcium/pharmacology Cell Membrane/enzymology Chromatography, Gel Cytosol/enzymology Deoxycholic Acid Humans In Vitro Techniques Phosphatidylinositol 4,5-Diphosphate Phosphatidylinositols/blood Type C Phospholipases/blood,isolation & purification
Chemicals
Phosphatidylinositol 4,5-Diphosphate Phosphatidylinositols Deoxycholic Acid Type C Phospholipases Calcium
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Banno Y
Department of Biochemistry, Gifu University School of Medicine, Japan.
Nozawa Y
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37 references, click to expand
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Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2480095
Published
1987-11-15
Pages
95-101
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1148504
Subset
IM
External Links
PubMed source DOI: 10.1042/bj2480095 PMC Full Text View journal details
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