Triton X-114: a detergent that has come in from the cold

Pryde, James G.

doi:10.1016/0968-0004(86)90132-5

Cited by 143 publications

(74 citation statements)

References 17 publications

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“…Because Triton X -114 extract remains single phase at 4˚C but separates into aqueous and detergent phase at 30˚C, extraction with the detergent is very useful to fractionate cellular components based on hydrophobicity of molecules (Pryde, 1986). However, our results showed that both wild type and the non-palmitoylated form of Gsα partitioned in the aqueous phase of Triton X-114 fractionate, demonstrating that palmitoylation on Gsα was not sufficient solely to determine the partition.…”

Section: Discussioncontrasting

confidence: 53%

Retainment of membrane binding capacity of non-palmitoylated Gs α mutants expressed in COS-1 cells

Yang

Cho²,

Bae³

et al. 1998

Exp Mol Med

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IntroductionHeterotrimeric guanine nucleotide binding regulatory proteins (G proteins) transduce extracellular signals such as hormones, neurotransmitters and photons into intracellular signals by receptor-effector coupling. When the G protein binds to the ligand-activated receptor, it replaces the bound GDP with GTP. The GTP-bound activated G protein regulates effectors such as enzymes and ion channels (Gilman, 1987;Kaziro et al., 1991; Neer, 1995). Because most of the receptors coupled to G proteins are integral proteins that have seven hydro-phobic transmembrane domains, the G proteins need to have a membrane binding capacity to receive signals from the membrane receptors. The α subunit of G protein is known to bind tightly to the cytoplasmic face of the plasma membrane without any membrane spanning hydrophobic domain. The β γ subunit complex has a high affinity for the membrane because of isoprenylation of the gamma subunit, and plays an important role in anchoring the α subunit of G proteins (Sternweis, 1986). However, fatty acylation with myristic acid and palmitoylation was found to be important for membrane binding by α subunits of the G protein family (Jones et al., 1990, Mumby, 1997. The reversible and dynamic palmitoylation had been claimed to be critical in membrane binding by the alpha s u b u n i t of the stimulatory G protein (Gsα) (Wedegaertner et al., 1993;Wedegaertner and Bourne, 1994), but it was also argued that this domain of Gsα was not essential for membrane binding from the finding that Gsα m u t a n t s deleted in this domain retained membrane binding capacity (Degtyarev et al. , 1993;Juhnn, 1993). Thus it is necessary to make clear the importance of this modification in the binding of Gsα.The present study was performed to evaluate the role of palmitoylation in membrane binding of Gsα. We constructed Gsα mutants by replacing the glycine-2, the cognate site of myristoylation in Giα with alanine, and cysteine-3, the site of palmitoylation with serine, and then analyzed the intracellular distribution of the mutant proteins expressed in COS-1 cells by quantitative EXPERIMENTAL and MOLECULAR MEDICINE, Vol. 30, No 4, 235-239, December 1998 Jung Abbreviations: G protein, guanine nucleotide-binding regulatory protein; Gi, adenylyl cyclase inhibitory G protein; Gs, adenylyl cyclase stimulatory G protein; Gsα, α subunit of Gs; Gα,α subunit of G protein ; Gβγ, βγ subunit of G protein Retainment of membrane binding capacity of nonpalmitoylated Gs mutants expressed in COS-1 cells

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Section: Discussioncontrasting

confidence: 53%

Retainment of membrane binding capacity of non-palmitoylated Gs α mutants expressed in COS-1 cells

Yang

Cho²,

Bae³

et al. 1998

Exp Mol Med

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“…The severa1 pellet washes and SUC gradient interphases were responsible for the protein loss. The NPA-binding activity partitioned preferentially in the lipid-rich phase, indicating that the NPA-binding protein was more hydrophobic than the majority of the PM protein (Pryde, 1986). Although some activity was found in the detergent-rich phase, none was detected in the aqueous phase.…”

Section: Hydrophobic Phase During Triton X-114 Partitioningmentioning

confidence: 93%

“…Although this partitioning is not a solubilization protocol per se (Pryde, 1986), it allows an approximate determination of the hydrophobicity of a protein and has been successfully used in animal (Pryde and Phillips, 1986) and plant systems. A recent example is the resolution of the membrane localization of the fusicoccin receptor in Commelina communis tissues (Oecking et al, 1994).…”

Section: Hydrophobic Phase During Triton X-114 Partitioningmentioning

confidence: 99%

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The N-1-Naphthylphthalamic Acid-Binding Protein Is an Integral Membrane Protein

Bernasconi¹,

Patel²,

Reagan³

et al. 1996

Plant Physiology

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N-1 -Naphthylphthalmic acid (NPA)-binding protein is a plasmalemma (PM) protein involved in the control of cellular auxin efflux. We re-evaluated the spatial relationship of this protein with the PM of zucchini (Cucurbifa pepo L.) hypocotyls. First, Triton X-114 partitioning indicated that the NPA-binding protein was more hydrophobic than most PM proteins. Second, the NPA-binding activity was found to be resistant to proteolytic digestion in membranes.

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“…In Triton X-114 phase partition it was unexpectedly partitioned into the aqueous phase, similar to the behaviour of peripheral membrane proteins [36], and in clear contrast to the also highly glycosylated LAMPs which were mainly partitioned into the detergent phase. However, some integral membrane glycoproteins are also known to anomalously partition into the aqueous phase, possibly because their large hydrophilic domains prevent them from being intercalated into the hydrophobic interior of the detergent micelle [47]. Therefore, it remains at present unresolved whether the teratocarcinoma glycoprotein is an integral or peripheral membrane component.…”

Section: Discussionmentioning

confidence: 99%

Identification of a major poly‐N‐acetyllactosamine‐containing cell‐surface glycoprotein of mouse teratocarcinoma cells

Spillmann

Finne

1994

European Journal of Biochemistry

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Mouse teratocarcinoma F9 cells were induced to primitive endoderm differentiation with retinoic acid, and poly-N-acetyllactosamine-containing surface glycoproteins were identified by radiolabelling endo-P-galactosidase-cleavable glycans with galactosyltransferase and radiolabelled UDPgalactose. One major radiolabelled band with an apparent size of 250-500 kDa was identified which differed from the known poly-N-acetyllactosamine-containing glycoproteins laminin, fibronectin, lysosome-associated membrane protein (LAMP)-1 and LAMP-2. This acidic glycoprotein, resistant to glycosaminoglycan-degrading enzymes and proteases, was purified by extraction and phase partition with Triton X-114, octyl Sepharose and Helix pomatiu lectin chromatography. The purified glycoprotein could be digested by endo-p-galactosidase and glycopeptide N-glycosidase F to an apparent size of 160-240 kDa. During retinoic-acid-induced differentiation into primitive endoderm cells, the glycoprotein showed a several-fold increase and a broadening to an apparent size of 200 -> 700 kDa. The glycoprotein was no longer detected in retinoic-acid and dibutyryl-CAMP-treated cells which had undergone further differentiation to parietal endoderm cells, nor in the permanently differentiated parietal endoderm line F9-AC. The results suggest that the glycoprotein is a major carrier of poly-N-acetyllactosamine chains on differentiating teratocarcinoma F9 cells, and that its expression as revealed by the poly-N-acetyllactosamine labelling method is regulated by the stage of cellular differentiation, After their discovery in erythrocytes and teratocarcinoma cells

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Triton X-114: a detergent that has come in from the cold

Cited by 143 publications

References 17 publications

Retainment of membrane binding capacity of non-palmitoylated Gs α mutants expressed in COS-1 cells

Retainment of membrane binding capacity of non-palmitoylated Gs α mutants expressed in COS-1 cells

The N-1-Naphthylphthalamic Acid-Binding Protein Is an Integral Membrane Protein

Identification of a major poly‐N‐acetyllactosamine‐containing cell‐surface glycoprotein of mouse teratocarcinoma cells

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