title: Activation of phospholipase C-beta 2 mutants by G protein alpha q and beta gamma subunits.
abstract: The beta- but not the gamma- and delta-type isozymes of inositol phospholipid-specific phospholipase C (PLC) are activated by G protein alpha q and beta gamma subunits. The beta-type PLC isozymes differ from other isozymes in that they contain a long carboxyl-terminal region downstream of the Y catalytic domain and a region rich in acidic amino acids between the two separated X and Y catalytic domains. To determine the sites on PLC-beta 2 that participate in the interaction of the enzyme with alpha q and beta gamma subunits, we introduced specific truncations and substitutions in the PLC-beta 2 cDNA at positions corresponding to the carboxyl-terminal and acidic amino acid-rich regions, respectively. After transient expression of these cDNA clones in CV-1 cells, the mutant enzymes were partially purified and their capacity to be activated by alpha q and beta gamma subunits determined. Substitution of glutamine residues for three or all seven of a stretch of consecutive glutamic acids in the acidic domain of PLC-beta 2 affected neither alpha q- nor beta gamma-dependent activation significantly. Carboxyl-terminal truncation to residue Gly-934 or to residue Ala-867 resulted in enzymes that were activated by beta gamma but not by alpha q. This result suggests that the carboxyl-terminal region of PLC-beta 2 is required for activation by alpha q, and that beta gamma subunits interact with a different region of the enzyme. Thus, alpha q and beta gamma subunits may independently modulate a single PLC-beta 2 molecule concurrently.
authors.affiliation: Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: S B
authors.initials: SB
authors.lastname: Lee
authors.forename: S H
authors.initials: SH
authors.lastname: Shin
authors.forename: J R
authors.initials: JR
authors.lastname: Hepler
authors.forename: A G
authors.initials: AG
authors.lastname: Gilman
authors.forename: S G
authors.initials: SG
authors.lastname: Rhee
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000494:pharmacology; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019269:Phosphatidylinositol 4,5-Diphosphate; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D014162:Transfection
chemical_list: D017931:DNA Primers; D007527:Isoenzymes; D046911:Macromolecular Substances; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D011994:Recombinant Proteins; D010727:Phosphoric Diester Hydrolases; D019204:GTP-Binding Proteins; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D002118:Calcium