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PTMS_gprotein.md

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Notes on G protein PTMs

Gα

Gαi is myristoylated and palmitoylated

According to Villaseca et al. 2022, Gαi "membrane location is mediated by two lipid modification, myristoylation and palmitoylation at the amino terminus. Gαi family is the only family that is myristoylated, in contrast with the other Gα (Gαs, Gα12/13, and Gαq) families that are only palmitoylated; both lipid modifications are important to allow interaction with the membrane, in order to interact with their receptor and with their membrane effectors Chen and Manning, 2001 Morales et al., 1998 Fishburn et al., 1999

PTMs sites on Gαi

  • Morales et al., 1998 reports that the double lipidation is established for all members of the αi family (αi1, αi2, αi3, αz, and αo) (myristoylated on the N-terminal glycine residue and palmitoylated on the adjacent cysteine).

  • Wedegaertner et al. 1995 is also in agreement with the double lipidation, and indicates the sites. Gαi should be N-myristoylation at G and palmitoylated at the C ("[...] attached through a labile, reversible thioester linkage to a cysteine near the N terminus")

  • Wedegaertner et al. 1995 also states that ''Myristoylation, or more specifically N-myristoylation, is the result of co-translational addition of the saturated 14-carbon fatty acid myristate to a glycine residue at the extreme N terminus after removal of the initiating methionine''. This is also reiterated in Vögler et al. 2008 "The absolute requirement for a free glycine residue at the N-terminus of the substrate protein implies that the initiating methionine must be eliminated by a methionyl amino peptidase prior to lipidation."

  • image from Wedegaertner

Gαi palmitoylation is reversible, and Gαi myristoylation is a precondition for Gi palmitoylation

  • Wedegaertner et al. 1995 as noted above, the palmitoylated thioester linkage is "reversible" and "labile" (though no timescales or further details are given). (Chen and Manning, 2001) not that ''The reversible nature of palmitoylation is intriguing in this regard, as it lends itself to a regulation integrated with the activation state of the G protein.''

  • Villaseca et al. 2022 It quickly became evident that, for α subunits of the Gi family, N-myristoylation was a prerequisite to palmitoylation (Mumby et al., 1994; Hallak et al., 1994a; Galbiati et al., 1994), and that the palmitoyl moiety alone (and certainly in conjunction with the N-myristoyl moiety), was sufficient for attachment of at least peptides to membrane (Shahinian and Silvius, 1995).

  • Vögler et al. 2008 For the Gi family myristoylation is a precondition for palmitoylation [32], [48], the secondary lipid modification found on Gα subunits. In this context, the role of the myristoyl group is to facilitate an initial contact with the membrane and to direct the Gα subunit to the palmitoylation compartment.

  • Vögler et al. 2008 also state that "the presence of a myristoyl moiety is not sufficient for stable membrane attachment RM Peitzsch 1993, [...] [but] mutations in Gα subunits that prevet N-myristoylation prevent protein binding to membranes Jones 1990, Mumby 1990". Therefore myristoyl group could "facilitate an initial contact with the membrane and to direct the Gα subunit to the palmitoylation compartment".

myristoylation can play a role in the structure of Gαi

  • The function of myristoylation could be "to maintain the N-terminal structure of the corresponding Gα subunit", as proposed by Hubbell and Hamm Preininger et al 2003: "To gain insight into the role of myristoylation in the structure and function of Gαi subunits, EPR and fluorescence studies have now been conducted using myristoylated Gαi proteins"

  • van Keulen and Rothlisberger, 2017 ran a 2 μs simulation of myristoylated Gi

Gγ1 and Gγ2 are prenylated at the C-terminal

  • Wedegaertner et al. 1995 G protein γ subunits are covalently modified by the 20-carbon isoprenoid geranylgeranyl or, (in the case of retinal-specific γ1, the 15-carbon isoprenoid farnesyl).

  • Also, Wedegaertner et al. 1995 mentions that "[...] the geranylgeranyl (or farnesyl) moiety is attached via a stable thioether bond to a cysteine residue located in the C-terminal “CAAX” box of γ . This is followed by proteolytic removal of the C-terminal three amino acids and then by carboxyl methylation at the new C terminus." and "After prenylation, the C-terminal three amino acids are removed, and the new C terminus is carboxylmethylated."

prenylation = geranylgeranylation or farnesylation

Gαs

  • Older papers (like Wedegaertner et al. 1993) state that Gs is only palmitoylated on Cys3,

  • Kleuss et al. 2003 determined [...] a second palmitoyl residue that is linked to the G protein at the extreme N-terminal glycine by an unusual amide linkage.

  • Kleuss et al. 2003 ''Under cellular conditions, Gαs most probably is temporarily palmitoylated on both residues, Gly2 and Cys3. The two neighbouring palmitoyl groups in Gαs are not equivalent, [...] The newly described palmitoyl group on Gly2 is attached via an amide linkage, while the palmitoyl group at Cys3 is attached via a thioester bond (Linder et al., 1993). So far, palmitoylation via an N-terminal glycine was unknown to occur on heterotrimeric G proteins. Furthermore, palmitoylation via an amide bond (N-terminal cysteine or internal lysine) seems to be rare.

  • Vögler et al. 2008 also reports "As an exception to the rule it was recently found that native Gαs can also be palmitoylated at the N-terminal glycine (Gly2), and that this lipid modification occurs via N-acylation Kleuss et al. 2003. The apparent dual Gly2/Cys3 palmitoylation of Gαs is similar to the myristoylation/palmitoylation motif of the Gαi family and it has functional consequences on signal transduction Kleuss and Gilman. "

force fields

Other refs

Related papers, but not directly addressing the issue of PTMs of G proteins