Mechanism of enzymatic reaction and protein-protein interactions of PLD from a 3D structural model.

Abstract

The phospholipase D (PLD) superfamily catalyzes the hydrolysis of cell membrane phospholipids generating the key intracellular lipid second messenger phosphatidic acid. However, there is not yet any resolved structure either from a crystallized protein or from NMR of any mammalian PLDs. We propose here a 3D model of the PLD2 by combining homology and ab initio 3 dimensional structural modeling methods, and docking conformation. This model is in agreement with the biochemical and physiological behavior of PLD in cells. For the lipase activity, the N- and C-terminal histidines of the HKD motifs (His 442/His 756) form a catalytic pocket, which accommodates phosphatidylcholine head group (but not phosphatidylethanolamine or phosphatidyl serine). The model explains the mechanism of the reaction catalysis, with nucleophilic attacks of His 442 and water, the latter aided by His 756. Further, the secondary structure regions superimposed with bacterial PLD crystal structure, which indicated an agreement with the model. It also explains protein-protein interactions, such as PLD2-Rac2 transmodulation (with a 1:2 stoichiometry) and PLD2 GEF activity both relevant for cell migration, as well as the existence of binding sites for phosphoinositides such as PIP2. These consist of R236/W238 and R557/W563 and a novel PIP2 binding site in the PH domain of PLD2, specifically R210/R212/W233. In each of these, the polar inositol ring is oriented towards the basic amino acid Arginine. Since tumor-aggravating properties have been found in mice overexpressing PLD2 enzyme, the 3D model of PLD2 will be also useful, to a large extent, in developing pharmaceuticals to modulate its in vivo activity.

• Authors: Madhu Mahankali, Gerald Alter, Julian Gomez-Cambronero
• Journal: Cellular signalling (Cell Signal) Vol. 27 Issue 1 Pg. 69-81 (Jan 2015) ISSN: 1873-3913 [Electronic] England
• PMID: 25308783 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2014. Published by Elsevier Inc.

Chemical References

• Amino Acids
• Guanine Nucleotide Exchange Factors
• Isoenzymes
• Phosphatidylcholines
• Phosphatidylinositol 4,5-Diphosphate
• phospholipase D2
• Phospholipase D
• rac GTP-Binding Proteins

Topics

• Amino Acid Sequence
• Amino Acids (metabolism)
• Animals
• Binding Sites
• COS Cells
• Chlorocebus aethiops
• Guanine Nucleotide Exchange Factors (metabolism)
• Humans
• Isoenzymes (chemistry, metabolism)
• Mice
• Models, Molecular
• Molecular Sequence Data
• Phosphatidylcholines (metabolism)
• Phosphatidylinositol 4,5-Diphosphate (metabolism)
• Phospholipase D (chemistry, metabolism)
• Protein Binding
• Sequence Alignment
• Software
• Substrate Specificity
• rac GTP-Binding Proteins (metabolism)