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

Jung Mee Yang, Chin Ho Cho, Chang Dae Bae, Yong Sung Juhnn

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Abstract

Heterotrimeric guanine nucleotide binding regulatory proteins (G proteins) transduce extracellular signals into intracellular signals by coupling receptors and effectors. Because most of the G protein-coupled receptors are integral proteins, the G proteins need to have a membrane binding capacity to receive signals from the receptors. The α subunit of G protein binds tightly to the cytoplasmic face of the plasma membrane without any membrane spanning domain. Fatty acylation of Gsα with myristic acid or palmitic acid, in addition to the βγ subunits, plays an important role in anchoring the Gsα subunit. The reversible and dynamic palmitoylation of the alpha subunit of stimulatory G protein (Gsα) has been suggested as essential for its membrane attachment. However, in our previous experiments, Gsα deleted in the amino terminus containing palmitoylation site, retained its binding capacity when expressed in COS cells. Thus, to evaluate the role of palmitoylation in Gsα membrane binding, we constructed and expressed non-palmitoylated mutants of Gsα and analyzed their subcellular distributions in COS-1 cells. We found that non-palmitoylated mutants of Gsα, C3S- and G2A/C3S Gsα, retained their membrane binding capacities in COS-1 cells, demonstrating that palmitoylation is not essential for membrane binding of Gsα in COS-1 cells. We also found that the palmitoylation did not change significantly the distribution of Gsα in Triton X-114 partition. These results suggest that the palmitoylation of Gsα may produce different effects on membrane binding depending on cell types.

Original languageEnglish
Pages (from-to)235-239
Number of pages5
JournalExperimental and Molecular Medicine
Volume30
Issue number4
DOIs
StatePublished - 31 Dec 1998

Keywords

  • G proteins
  • Immunoblot
  • Membrane binding
  • Palmitoylation
  • Site-directed mutagenesis
  • Transfection
  • Triton X-114

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