title: Binding of phosphoinositide-specific phospholipase C-zeta (PLC-zeta) to phospholipid membranes: potential role of an unstructured cluster of basic residues.
abstract: Phospholipase C-zeta (PLC-zeta) is a sperm-specific enzyme that initiates the Ca2+ oscillations in mammalian eggs that activate embryo development. It shares considerable sequence homology with PLC-delta1, but lacks the PH domain that anchors PLC-delta1 to phosphatidylinositol 4,5-bisphosphate, PIP2. Thus it is unclear how PLC-zeta interacts with membranes. The linker region between the X and Y catalytic domains of PLC-zeta, however, contains a cluster of basic residues not present in PLC-delta1. Application of electrostatic theory to a homology model of PLC-zeta suggests this basic cluster could interact with acidic lipids. We measured the binding of catalytically competent mouse PLC-zeta to phospholipid vesicles: for 2:1 phosphatidylcholine/phosphatidylserine (PC/PS) vesicles, the molar partition coefficient, K, is too weak to be of physiological significance. Incorporating 1% PIP2 into the 2:1 PC/PS vesicles increases K about 10-fold, to 5x10(3) M-1, a biologically relevant value. Expressed fragments corresponding to the PLC-zeta X-Y linker region also bind with higher affinity to polyvalent than monovalent phosphoinositides on nitrocellulose filters. A peptide corresponding to the basic cluster (charge=+7) within the linker region, PLC-zeta-(374-385), binds to PC/PS vesicles with higher affinity than PLC-zeta, but its binding is less sensitive to incorporating PIP2. The acidic residues flanking this basic cluster in PLC-zeta may account for both these phenomena. FRET experiments suggest the basic cluster could not only anchor the protein to the membrane, but also enhance the local concentration of PIP2 adjacent to the catalytic domain.
authors: Cell Signaling Laboratory, Wales Heart Research Institute, UK.
authors: Michail
authors: M
authors: Nomikos
authors: Anna
authors: A
authors: Mulgrew-Nesbitt
authors: Payal
authors: P
authors: Pallavi
authors: Gyongyi
authors: G
authors: Mihalyne
authors: Irina
authors: I
authors: Zaitseva
authors: Karl
authors: K
authors: Swann
authors: F Anthony
authors: FA
authors: Lai
authors: Diana
authors: D
authors: Murray
authors: Stuart
authors: S
authors: McLaughlin
mesh_terms: D000818:Animals; D020134:Catalytic Domain; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism; D008567:Membranes, Artificial; D051379:Mice; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000737:chemistry* / Q000378:metabolism; D054801:Phosphoinositide Phospholipase C; D054803:Phospholipase C delta; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity; D010738:Type C Phospholipases / Q000737:chemistry* / Q000378:metabolism
chemical_list: D007527:Isoenzymes; D008567:Membranes, Artificial; D010455:Peptides; D019269:Phosphatidylinositol 4,5-Diphosphate; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D054803:Phospholipase C delta; C518025:Plcd1 protein, mouse; C518026:Plcd1 protein, rat; C499234:Plcz1 protein, mouse