Anionic phospholipids,annexins and the activity of phospholipases A2
Anionic phospholipids,annexins and the activity of phospholipases A2
The effects of annexin V and anionic phospholipids on PLA2 activity has been studied, primarily using a continuous, fluorescence displacement assay.
The annexins are known to inhibit phospholid hydrolysis by secretory PLA2's (sPLA2) and a mechanism of substrate depletion is proposed. Using annexin V, a mechanism of substrate depletion was confirmed for the inhibition of various sPLA2's against phospholipid substrates. Moreover, the results suggest the term interfacial competition best describes the inhibitory effect and as such provided a means of assessing the relative affinities of sPLA2's for anionic phospholipids. The results highlight the very high affinity of the human non-pancreatic sPLA2 (human sPLA2) for such vesicles while Naja naja sPLA2 (nn-sPLA2) and porcine pancreatic sPLA2 (pp-sPLA2) showed lower affinities. Hydrolysis of vesicles containing phosphatidylserine and phosphatidylcholine (PC) were differentially inhibited by annexin V. The differences must reflect the preference of annexin V and pp and human sPLA2 for anionic phospholipids, while nn-SPLA2 is readily able to hydrolyse PC interfaces. Studies with another anionic phospholipid, cardiolipin, and pp-sPLA2, showed inhibition was more complex than with other phospholipids tested.
In contrast to sPLA2 enzymes, studies involving annexins and cytosolic PLA2 (cPLA2) are very limited. In this study annexin V was shown to inhibit cPLA2 activity by substrate depletion, while this inhibition was dependent on the nature of the phospholipid and the calcium concentration. Hydrolysis of PC substrate by cPLA2 was not significantly affected by annexin V over a range of calcium concentrations, presumably reflecting the inability of annexin V to bind zwitterionic interfaces. In contrast, hydrolysis of anionic substrates was readily inhibited by annexin V. However, the calcium concentrations required are greater than those associated with calcium stimulated events, and with cPLA2 activity within the cell, questioning the potential of annexin V in regulating cPLA2 activity.
Phosphatidic acid was found to enhance cPLA2 activity and could be generated within a phospholipid interface by phospholipase D. Such treatment appeared to make phospholipd vesicles more amenable to hydrolysis by cPLA2.
University of Southampton
Buckland, Andrew.G
c0b3a024-9df7-4414-9853-11fc6e55f7d1
1998
Buckland, Andrew.G
c0b3a024-9df7-4414-9853-11fc6e55f7d1
Buckland, Andrew.G
(1998)
Anionic phospholipids,annexins and the activity of phospholipases A2.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
The effects of annexin V and anionic phospholipids on PLA2 activity has been studied, primarily using a continuous, fluorescence displacement assay.
The annexins are known to inhibit phospholid hydrolysis by secretory PLA2's (sPLA2) and a mechanism of substrate depletion is proposed. Using annexin V, a mechanism of substrate depletion was confirmed for the inhibition of various sPLA2's against phospholipid substrates. Moreover, the results suggest the term interfacial competition best describes the inhibitory effect and as such provided a means of assessing the relative affinities of sPLA2's for anionic phospholipids. The results highlight the very high affinity of the human non-pancreatic sPLA2 (human sPLA2) for such vesicles while Naja naja sPLA2 (nn-sPLA2) and porcine pancreatic sPLA2 (pp-sPLA2) showed lower affinities. Hydrolysis of vesicles containing phosphatidylserine and phosphatidylcholine (PC) were differentially inhibited by annexin V. The differences must reflect the preference of annexin V and pp and human sPLA2 for anionic phospholipids, while nn-SPLA2 is readily able to hydrolyse PC interfaces. Studies with another anionic phospholipid, cardiolipin, and pp-sPLA2, showed inhibition was more complex than with other phospholipids tested.
In contrast to sPLA2 enzymes, studies involving annexins and cytosolic PLA2 (cPLA2) are very limited. In this study annexin V was shown to inhibit cPLA2 activity by substrate depletion, while this inhibition was dependent on the nature of the phospholipid and the calcium concentration. Hydrolysis of PC substrate by cPLA2 was not significantly affected by annexin V over a range of calcium concentrations, presumably reflecting the inability of annexin V to bind zwitterionic interfaces. In contrast, hydrolysis of anionic substrates was readily inhibited by annexin V. However, the calcium concentrations required are greater than those associated with calcium stimulated events, and with cPLA2 activity within the cell, questioning the potential of annexin V in regulating cPLA2 activity.
Phosphatidic acid was found to enhance cPLA2 activity and could be generated within a phospholipid interface by phospholipase D. Such treatment appeared to make phospholipd vesicles more amenable to hydrolysis by cPLA2.
This record has no associated files available for download.
More information
Published date: 1998
Identifiers
Local EPrints ID: 463321
URI: http://eprints.soton.ac.uk/id/eprint/463321
PURE UUID: 97c795b5-ce43-4c95-b18a-f3171a189d16
Catalogue record
Date deposited: 04 Jul 2022 20:50
Last modified: 04 Jul 2022 20:50
Export record
Contributors
Author:
Andrew.G Buckland
Download statistics
Downloads from ePrints over the past year. Other digital versions may also be available to download e.g. from the publisher's website.
View more statistics