Publications
211 results found
Parenti M, Magee AI, 1995, Fatty acid- and isoprenoid-linked membrane proteins, Biomembranes: A Multi-Volume Treatise, Vol: 1, Pages: 79-105, ISSN: 1874-5342
This chapter discusses the major structural, enzymological, and functional features of acylation with the fatty acids myristate and palmitate, and of isoprenylation and its associated modifications. Protein acylation refers to either co-translational or post-translational modifications that result in the covalent attachment of a fatty acid prosthetic group onto an acceptor protein. Two types of long chain saturated fatty acids serve as major substrates for these modifications: the 14-carbon myristic and the 16-carbon palmitic acids. Less commonly, several other fatty acids (such as stearic or oleic acids) have been found linked to proteins, and there are also interconversion reactions that occur after the fatty acid has been attached to the protein. Myristoylation is catalyzed by the soluble enzyme myristoyl-CoA: protein N-myristoyl transferase (NMT). The enzyme was originally purified from Saccharomyces cerevisiae and its complete sequence is known—it is a 455-residue monomeric protein located in the cytoplasm and not detectably associated with membranes of any cellular organelles. Palmitoylation of transmembrane proteins takes place after the translation of the respective polypeptide and before the trimming of peripheral mannose residues, a putative cis-Golgi function, has been completed. Palmitoylation sites for integral membrane proteins are usually close to the cytoplasmic surface of the transmembrane regions, suggesting that the active site of the enzyme is cytoplasmically exposed. © 1995, Elsevier Inc.
Milligan G, Parenti M, Magee AI, 1995, Erratum: The dynamic role of palmitoylation in signal transduction (TIBS 20, 181-186 (May 1995)), Trends in Biochemical Sciences, Vol: 20, ISSN: 0968-0004
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ANGST BD, BUXTON RS, MAGEE AI, 1995, CHARACTERIZATION OF HUMAN CARDIAC DESMOSOMAL CADHERINS, CARDIAC GROWTH AND REGENERATION, Vol: 752, Pages: 101-104, ISSN: 0077-8923
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MAGEE AI, WOOTON J, DEBONY J, 1995, DETECTING RADIOLABELED LIPID-MODIFIED PROTEINS IN POLYACRYLAMIDE GELS, LIPID MODIFICATIONS OF PROTEINS, Vol: 250, Pages: 330-336, ISSN: 0076-6879
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- Citations: 24
Koegl M, Zlatkine P, Ley SC, et al., 1994, Palmitoylation of multiple Src-family kinases at a homologous N-terminal motif., Biochem J, Vol: 303 ( Pt 3), Pages: 749-753, ISSN: 0264-6021
We have recently identified a novel N-terminal cysteine-containing motif which specifies the palmitoylation of several G-protein alpha-subunits [Parenti, Viganó, Newman, Milligan and Magee (1993) Biochem. J. 291, 349-353]. A related motif occurs at the N-terminus of members of the Src family of protein tyrosine kinases except for Src itself and Blk. We have investigated whether the Src, Fyn, Yes and Lck gene products are palmitoylated. Src was not labelled with [3H]palmitate when endogenously expressed in COS cells. In contrast, endogenous Yes immunoprecipitated from COS cells was palmitoylated. Fyn was palmitoylated in insect cells infected with a recombinant baculovirus and the palmitoylation was independent of protein synthesis, suggesting a dynamic turnover of this lipid. Fatty acid analysis indicated that most of the label was incorporated as palmitate. Lck was palmitoylated when expressed by transfection in COS cells. All of these protein tyrosine kinases were also detectably myristoylated in each of the systems tested. Experiments performed with mutants of Lck expressed by transfection in COS cells indicated that cysteines at positions 3 and 5 were both palmitoylation sites and that myristoylation was required for palmitoylation. To confirm that palmitoylation was occurring on cysteines in the N-terminal region of Fyn, site-directed mutagenesis was used to replace the cysteines at positions 3 and 6 with alanine. The resulting protein was not palmitoylated but was still myristoylated when expressed in COS cells. A glycine to alanine mutant at position 2 was also not palmitoylated, showing that myristoylation is a prerequisite for palmitoylation. Our data indicate that Src family members containing the N-terminal cysteine motif are indeed palmitoylated. By analogy with Ras, it is possible that palmitoylation may play an important role in the localization and function of Src family protein tyrosine kinases.
GALBIATI F, GUZZI F, MAGEE AI, et al., 1994, N-TERMINAL FATTY ACYLATION OF THE ALPHA-SUBUNIT OF THE G-PROTEIN G(I)1 - ONLY THE MYRISTOYLATED PROTEIN IS A SUBSTRATE FOR PALMITOYLATION, BIOCHEMICAL JOURNAL, Vol: 303, Pages: 697-700, ISSN: 0264-6021
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- Citations: 46
KOEGL M, ZLATKINE P, LEY SC, et al., 1994, PALMITOYLATION OF MULTIPLE SRC-FAMILY KINASES AT A HOMOLOGOUS N-TERMINAL MOTIF, BIOCHEMICAL JOURNAL, Vol: 303, Pages: 749-753, ISSN: 0264-6021
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- Citations: 135
Galbiati F, Guzzi F, Magee AI, et al., 1994, N-terminal fatty acylation of the alpha-subunit of the G-protein Gi1: only the myristoylated protein is a substrate for palmitoylation., Biochem J, Vol: 303 ( Pt 3), Pages: 697-700, ISSN: 0264-6021
The alpha-subunit of the G-protein Gi1 carries two fatty acyl moieties covalently bound to its N-terminal region: myristic acid is linked to glycine-2 and palmitic acid is linked to cysteine-3. Using site-directed mutagenesis on a cDNA construct of alpha i1 we have generated an alpha i1-G2A mutant, carrying alanine instead of glycine at position 2, and alpha i1-C3S mutant, in which serine replaced cysteine-3 and a double mutant with both substitutions (alpha i1-G2A/C3S). These constructs were individually expressed by transfection in Cos-7 cells, and incorporation of fatty acids into the various mutants was compared with wild-type alpha i1 monitoring metabolic labelling with [3H]palmitate or [3H]myristate. The disruption of the palmitoylation site in alpha i1-C3S did not influence myristoylation, whereas prevention of myristoylation in alpha i1-G2A also abolished palmitoylation. Co-translational myristoylation is thus an absolute requirement for alpha i1 to be post-translationally palmitoylated. The non-palmitoylated alpha i1-C3S showed reduced membrane binding to the same extent as the non-myristoylated/non-palmitoylated alpha i1-G2A and alpha i1-G2A/C3S mutants, indicating that the attachment of palmitic acid is necessary for proper interaction with the membrane.
GRASSIE MA, MCCALLUM JF, GUZZI F, et al., 1994, THE PALMITOYLATION STATUS OF THE G-PROTEIN G(0)1-ALPHA REGULATES ITS AVIDITY OF INTERACTION WITH THE PLASMA-MEMBRANE, BIOCHEMICAL JOURNAL, Vol: 302, Pages: 913-920, ISSN: 0264-6021
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- Citations: 38
Grassie MA, McCallum JF, Guzzi F, et al., 1994, The palmitoylation status of the G-protein G(o)1 alpha regulates its activity of interaction with the plasma membrane., Biochem J, Vol: 302 ( Pt 3), Pages: 913-920, ISSN: 0264-6021
Plasmids containing cDNAs encoding either the wild-type guanine-nucleotide-binding protein G(o)1 alpha or the palmitoylation-negative cysteine-3-to-serine (C3S) mutant of G(o)1 alpha were transfected into Rat 1 cells, and clones stably expressing immunoreactivity corresponding to these polypeptides were isolated. Clones C5B (expressing wild-type G(o)1 alpha) and D3 (expressing the mutant form) were selected for detailed study. Immunoprecipitation of whole cell lysates of each clone labelled with either [3H]palmitate or [3H]myristate demonstrated incorporation of [3H]myristate into both wild-type and the C3S mutant of G(o)1 alpha, but that incorporation of hydroxylamine-sensitive [3H]palmitate was restricted to the wild type. When membrane and cytoplasmic fractions were prepared from cells of either the C5B or D3 clones, although immunodetection of wild-type G(o)1 alpha was observed only in the membrane fraction, the C3S mutant was present in both membrane and cytoplasmic fractions. Furthermore, a significant proportion of the C3S G(o)1 alpha immunoreactivity was also detected in the cytoplasmic fraction if immunoprecipitation of recently synthesized G(o)1 alpha was performed from fractions derived from cells pulse-labelled with [35S]Trans label. Pretreatment of cells of both clones C5B and D3 with pertussis toxin led to complete ADP-ribosylation of the cellular population of G(o)1 alpha in both cell types, irrespective of whether the polypeptide was subsequently found in the membrane or cytoplasmic fraction following cellular disruption. By contrast, separation of membrane and cytoplasmic fractions before pertussis-toxin-catalysed [32P]ADP-ribosylation allowed modification only of the membrane-associated G(o)1 alpha (whether wild-type or the C3S mutant). This labelling was decreased substantially by incubation of the membranes with guanosine 5'-[beta gamma-imido]triphosphate. No cytoplasmic G-protein beta subunit was detected immunologically, and the non-membrane-as
GIANNAKOUROS T, NEWMAN CMH, CRAIGHEAD MW, et al., 1993, POSTTRANSLATIONAL PROCESSING OF SCHIZOSACCHAROMYCES-POMBE YPT5-PROTEIN - IN-VITRO AND IN-VIVO ANALYSIS OF PROCESSING MUTANTS, JOURNAL OF BIOLOGICAL CHEMISTRY, Vol: 268, Pages: 24467-24474, ISSN: 0021-9258
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- Citations: 19
Grassie MA, McCallum JF, Parenti M, et al., 1993, Lack of N terminal palmitoylation of G protein alpha subunits reduces membrane association., Biochem Soc Trans, Vol: 21, ISSN: 0300-5127
NEWMAN CMH, MAGEE AI, 1993, POSTTRANSLATIONAL PROCESSING OF THE RAS SUPERFAMILY OF SMALL GTP-BINDING PROTEINS, BIOCHIMICA ET BIOPHYSICA ACTA, Vol: 1155, Pages: 79-96, ISSN: 0006-3002
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- Citations: 99
GIANNAKOUROS T, NEWMAN CMH, ARMSTRONG J, et al., 1993, PROCESSING OF THE SMALL GTP-BINDING PROTEIN SPYPT1P IN SCHIZOSACCHAROMYCES-POMBE AND IN MAMMALIAN COS CELLS, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, Vol: 192, Pages: 983-990, ISSN: 0006-291X
BUXTON RS, COWIN P, FRANKE WW, et al., 1993, NOMENCLATURE OF THE DESMOSOMAL CADHERINS, JOURNAL OF CELL BIOLOGY, Vol: 121, Pages: 481-483, ISSN: 0021-9525
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- Citations: 265
PARENTI M, VIGANO MA, NEWMAN CMH, et al., 1993, A NOVEL N-TERMINAL MOTIF FOR PALMITOYLATION OF G-PROTEIN ALPHA-SUBUNITS, BIOCHEMICAL JOURNAL, Vol: 291, Pages: 349-353, ISSN: 0264-6021
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- Citations: 153
Parenti M, Viganó MA, Newman CM, et al., 1993, A novel N-terminal motif for palmitoylation of G-protein alpha subunits., Biochem J, Vol: 291 ( Pt 2), Pages: 349-353, ISSN: 0264-6021
We have examined the post-translational processing of G alpha subunits expressed endogenously in rat PC12 and NG108-15 rat/mouse hybrid cells, and after transfection of cDNA expression constructs into COS cells. Thioester-linked palmitoylation of alpha o, alpha s, alpha q/alpha 11 and alpha 12 has been detected by metabolic labelling with [3H]palmitate and immunoprecipitation. Palmitoylation of alpha o occurs post-translationally in cells treated with protein-synthesis inhibitors, suggesting possible dynamic acylation. Palmitoylation of the C-terminal CAAX motif has been excluded. Site-directed mutagenesis of alpha o has been used to implicate the site of modification as a cysteine residue next to the N-terminal myristoylated glycine, in a novel protein-lipid modification motif Met-Gly-Cys. The non-palmitoylated alpha o mutant is still myristoylated but shows reduced membrane binding, suggesting that reversible palmitoylation may regulate G alpha localization and function.
KING IA, TABIOWO A, PURKIS P, et al., 1993, EXPRESSION OF DISTINCT DESMOCOLLIN ISOFORMS IN HUMAN EPIDERMIS, JOURNAL OF INVESTIGATIVE DERMATOLOGY, Vol: 100, Pages: 373-379, ISSN: 0022-202X
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- Citations: 33
ARNEMANN J, SULLIVAN KH, MAGEE AI, et al., 1993, STRATIFICATION-RELATED EXPRESSION OF ISOFORMS OF THE DESMOSOMAL CADHERINS IN HUMAN EPIDERMIS, JOURNAL OF CELL SCIENCE, Vol: 104, Pages: 741-750, ISSN: 0021-9533
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- Citations: 163
Arnemann J, Sullivan KH, Magee AI, et al., 1993, Stratification-related expression of isoforms of the desmosomal cadherins in human epidermis., J Cell Sci, Vol: 104 ( Pt 3), Pages: 741-750, ISSN: 0021-9533
Desmosomal junctions are abundant in epidermis and contain two classes of transmembrane glycoprotein, the desmocollins and the desmogleins, which are members of the cadherin superfamily of Ca(2+)-dependent cell adhesion molecules. The desmocollin subfamily includes DGIV/V and DGII/III while the desmoglein subfamily includes DGI, HDGC and the autoantigen of the blistering skin disease pemphigus vulgaris (PVA). There are also several non-glycosylated proteins, including the desmoplakins and plakoglobin, present in the desmosomal plaque, which forms a link between the glycoproteins and the cytokeratin intermediate filaments. To provide a picture of the expression of the desmosomal genes and their products in epidermis, we have used in situ hybridisation and immunofluorescence staining on sections of human foreskin. We find that, as expected, desmoplakin DPI/II and plakoglobin are expressed throughout the epidermis, gradually accumulating during differentiation, which probably reflects the increased numbers of desmosomes. In contrast, while keratin 14 and the hemidesmosomal component bullous pemphigoid antigen I (BPAGI) are basal-specific, desmocollin DGIV/V is expressed only in the upper spinous/granular layers of the epidermis, whereas DGII/III expression is enriched in the basal layers. Amongst the desmogleins, expression of DGI appears similar to desmoplakin and plakoglobin; PVA is more prevalent in the lower spinous layers, whereas HDGC expression is detected basally but not suprabasally. The major desmosomal cadherin transcripts are desmocollin DGIV/V and desmoglein DGI. The resultant changes in desmosomal composition and structure may reflect the maturation of desmosomes, presumably being related to the need for changes in cell adhesion during stratification, terminal differentiation, and desquamation, and point to the desmosome being a key player in epidermal differentiation.
Magee T, Newman C, 1992, The role of lipid anchors for small G proteins in membrane trafficking., Trends Cell Biol, Vol: 2, Pages: 318-323, ISSN: 0962-8924
Small GTP-binding proteins of the Ras superfamily play diverse roles in intracellular trafficking. In order to perform these functions, the proteins must associate with specific donor vesicles and be recycled after fusion of these vesicles with their acceptor membrane target. Recent results have identified a number of lipid modifications of these proteins, occurring at the N- or C-termini, that contribute to their membrane binding. Recycling appears, in some cases, to be mediated by soluble proteins that bind the lipid-modified tails, removing them from the membrane and allowing their reutilization via the cytosol.
NEWMAN CMH, GIANNAKOUROS T, HANCOCK JF, et al., 1992, POSTTRANSLATIONAL PROCESSING OF SCHIZOSACCHAROMYCES-POMBE YPT PROTEINS, JOURNAL OF BIOLOGICAL CHEMISTRY, Vol: 267, Pages: 11329-11336, ISSN: 0021-9258
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- Citations: 55
ARNEMANN J, SPURR NK, MAGEE AI, et al., 1992, THE HUMAN GENE (DSG2) CODING FOR HDGC, A SECOND MEMBER OF THE DESMOGLEIN SUBFAMILY OF THE DESMOSOMAL CADHERINS, IS, LIKE DSG1 CODING FOR DESMOGLEIN DGI, ASSIGNED TO CHROMOSOME-18, GENOMICS, Vol: 13, Pages: 484-486, ISSN: 0888-7543
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- Citations: 38
Buxton RS, Magee AI, 1992, Structure and interactions of desmosomal and other cadherins., Semin Cell Biol, Vol: 3, Pages: 157-167, ISSN: 1043-4682
The cadherin superfamily of cell-cell adhesion molecules is now known to include proteins of the desmosome as well as of the adherens type of junction. The desmosomal cadherins consist of two families of proteins, the desmocollins and the desmogleins, both of which are represented by different isoforms which are differentially expressed in epidermis. The desmocollins are quite similar to the classic cadherins in overall structure, but with alternatively spliced variants; the desmogleins have extra cytoplasmic sequences added onto the basic cadherin structure. The cytoplasmic domains are specialized for binding to 'mediator' proteins, such as plakoglobin, which interconnect to the intermediate filament system rather than the actin filaments as do the classic cadherins.
MAGEE AI, NEWMAN CMH, GIANNAKOUROS T, et al., 1992, LIPID MODIFICATIONS AND FUNCTION OF THE RAS SUPERFAMILY OF PROTEINS, BIOCHEMICAL SOCIETY TRANSACTIONS, Vol: 20, Pages: 497-499, ISSN: 0300-5127
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- Citations: 33
GIANNAKOUROS T, ARMSTRONG J, MAGEE AI, 1992, PROTEIN PRENYLATION IN SCHIZOSACCHAROMYCES-POMBE, FEBS LETTERS, Vol: 297, Pages: 103-106, ISSN: 1873-3468
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- Citations: 15
MAGEE AI, HANCOCK JF, FAWELL E, et al., 1992, THE RAS SUPERFAMILY - POSTTRANSLATIONAL MODIFICATIONS AND FUNCTIONAL REGULATION, INTERNATIONAL SYMP ON BIOTECHNOLOGY OF GROWTH FACTORS : VASCULAR AND NERVOUS SYSTEMS, Publisher: KARGER, Pages: 1-5
WHEELER GN, BUXTON RS, PARKER AE, et al., 1991, DESMOSOMAL GLYCOPROTEINS-I, GLYCOPROTEINS-II AND GLYCOPROTEINS-III - NOVEL MEMBERS OF THE CADHERIN SUPERFAMILY, Publisher: PORTLAND PRESS LTD, Pages: 1060-1064, ISSN: 0300-5127
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- Citations: 27
Wheeler GN, Buxton RS, Parker AE, et al., 1991, Desmosomal glycoproteins I, II and III: novel members of the cadherin superfamily., Biochem Soc Trans, Vol: 19, Pages: 1060-1064, ISSN: 0300-5127
Magee AI, Buxton RS, 1991, Transmembrane molecular assemblies regulated by the greater cadherin family, CURRENT OPINION IN CELL BIOLOGY, Vol: 3, Pages: 854-861, ISSN: 0955-0674
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- Citations: 132
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