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A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface.

Publication ,  Journal Article
Burke, JR; Gregor, KR; Padmanabha, R; Banville, J; Witmer, MR; Davern, LB; Manly, SP; Tramposch, KM
Published in: J Enzyme Inhib
June 1998

Cytosolic phospholipase A2 (cPLA2) catalyzes the selective release of arachidonic acid from the sn-2 position of phospholipids and is believed to play a key cellular role in the generation of arachidonic acid. When assaying the human recombinant cPLA2 using membranes isolated from [3H]arachidonate-labeled U937 cells as substrate, 3,3-Dimethyl-6-(3-lauroylureido)-7-oxo-4-thia-1-azabicyclo[3,2,0] heptane-2-carboxylic acid (1) was found to inhibit the enzyme in a dose-dependent manner (IC50 = 72 microM). This beta-lactam did not inhibit other phospholipases, including the human nonpancreatic secreted phospholipase A2. The inhibition of cPLA2 was found not to be time-dependent. This, along with the observation that the degradation of the inhibitor was not catalyzed by the enzyme, demonstrates that the inhibition does not result from the formation of an acyl-enzyme intermediate with the active site serine residue. Moreover, the ring-opened form of 1 is also able to inhibit cPLA2 with near-equal potency. To further characterize the mechanism of inhibition, an assay in which the enzyme is bound to vesicles of 1,2-dimyristoyl-sn-glycero-3-phosphomethanol containing 6-10 mole percent of 1-palmitoyl-2-[1-14C]-arachidonoyl-sn-glycero-3-phosphocholine was employed. With this substrate system, the dose-dependent inhibition was defined by kinetic equations describing competitive inhibition at the lipid/water interface. The apparent dissociation constant for the inhibitor bound to the enzyme at the interface (KI*app) was determined to be 0.5 +/- 0.1 mole% versus an apparent dissociation constant for the arachidonate-containing phospholipid of 0.4 +/- 0.1 mole%. Thus, 1 represents a novel structural class of inhibitors of cPLA2 which partitions into the phospholipid bilayer and competes with the phospholipid substrate for the active site.

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Published In

J Enzyme Inhib

DOI

ISSN

8755-5093

Publication Date

June 1998

Volume

13

Issue

3

Start / End Page

195 / 206

Location

Switzerland

Related Subject Headings

  • Water
  • Recombinant Proteins
  • Phospholipids
  • Phospholipases A2
  • Phospholipases A
  • Penicillanic Acid
  • Medicinal & Biomolecular Chemistry
  • Kinetics
  • Isoenzymes
  • Humans
 

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Burke, J. R., Gregor, K. R., Padmanabha, R., Banville, J., Witmer, M. R., Davern, L. B., … Tramposch, K. M. (1998). A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface. J Enzyme Inhib, 13(3), 195–206. https://doi.org/10.3109/14756369809028340
Burke, J. R., K. R. Gregor, R. Padmanabha, J. Banville, M. R. Witmer, L. B. Davern, S. P. Manly, and K. M. Tramposch. “A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface.J Enzyme Inhib 13, no. 3 (June 1998): 195–206. https://doi.org/10.3109/14756369809028340.
Burke JR, Gregor KR, Padmanabha R, Banville J, Witmer MR, Davern LB, et al. A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface. J Enzyme Inhib. 1998 Jun;13(3):195–206.
Burke, J. R., et al. “A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface.J Enzyme Inhib, vol. 13, no. 3, June 1998, pp. 195–206. Pubmed, doi:10.3109/14756369809028340.
Burke JR, Gregor KR, Padmanabha R, Banville J, Witmer MR, Davern LB, Manly SP, Tramposch KM. A beta-lactam inhibitor of cytosolic phospholipase A2 which acts in a competitive, reversible manner at the lipid/water interface. J Enzyme Inhib. 1998 Jun;13(3):195–206.
Journal cover image

Published In

J Enzyme Inhib

DOI

ISSN

8755-5093

Publication Date

June 1998

Volume

13

Issue

3

Start / End Page

195 / 206

Location

Switzerland

Related Subject Headings

  • Water
  • Recombinant Proteins
  • Phospholipids
  • Phospholipases A2
  • Phospholipases A
  • Penicillanic Acid
  • Medicinal & Biomolecular Chemistry
  • Kinetics
  • Isoenzymes
  • Humans