G Protein Trafficking - GPCR Signalling Complexes: Synthesis, Assembly, Trafficking and Specificity - page 189

co-translationally, more recently it has become clear that myristoylation can occur
post-translationally as well, usually when an internal glycine is exposed and
becomes an N-terminal amino acid after cleavage by the apoptotic caspases
(Martin et al. 2011 ). So far, there is no evidence to indicate that Ga i can be post-
translationally myristoylated.
Myristate is not an abundant fatty acid in the cell, and thus the myristoylation
process is very selective for myristate as opposed to other more abundant fatty acids
(reviewed recently in Martin et al. 2011 ) . However, the retinal-speci fi c G a t , as well
as several other retinal proteins, has been found to be heterogeneously acylated at
the N-terminal glycine (Kokame et al. 1992 ; Johnson et al. 1994 ; Neubert et al.
1992 ). Instead of only having myristate (C14:0, in which C14 indicates the carbon
chain length, and 0 indicates the number of unsaturated bonds) attached, Ga t also
exists in isoforms with attached 5-cis-tetradecenoic acid (C14:1), 5-cis, 8-cis-
tetradecadienoic acid (C14:2) and laurate (C12). This heterogeneous N-acylation
appears to occur only in the retinal photoreceptor cells due to unique characteristics
of lipid metabolism in the retina compared to other cells in the body (Bereta and
Palczewski 2011 ). The function of the differently modified isoforms of Ga t remains
incompletely understood. Studies have indicated that laurate modified forms bind
with lower affinity to Gb g and have lower steady-state GTPase activity (Hashimoto
et al. 2004 ; Kokame et al. 1992 ), suggesting that the lipid moiety can influence
protein-protein interactions or intramolecular interactions. Alternatively, the func-
tional differences may simply reflect that Ga t isoforms with less hydrophobic fatty
acids attached have lower affinity for membranes and thus decreased function.
Regardless, the existence of differently modified forms of Ga t provides the oppor-
tunity for important functional differences, and it remains possible that different
isoforms of Ga t with different affinities for membranes could exhibit differences in
trafficking. This speculation remains to be examined.
Lastly, myristoylation appears to be an irreversible modification. Since myristate
is added co-translationally and mechanisms for its removal do not seem to exist,
myristate appears to be attached to Ga i subunits throughout their lifetime; thus,
myristoylation does not fit the concept of a regulatory modification. However, the
relatively weak affinity of myristate for membranes, allows other factors to regulate
the reversible membrane binding of myristoylated proteins (Resh 2006 ) . For exam-
ple, the covalent attachment of another lipid, such as palmitate (discussed below)
can synergize with myristate to provide very strong membrane binding, while phos-
phorylation of a myristoylated protein can shift the protein off of the membrane by
providing electrostatic repulsion from acidic membrane lipid headgroups. In this
way covalently attached myristate, although irreversible, can affect membrane
localization and trafficking of proteins, such as Ga i subunits.
In contrast to myristoylation, palmitoylation occurs post-translationally, is read-
ily reversible, lacks a clear consensus motif and functions as a stronger membrane
anchor due to its greater hydrophobicity. With the exception of Ga t and Ga gust , all
Ga subunits can be palmitoylated at one or more cysteines located within the first
20 N-terminal amino acids (Table 11.1 ). For members of the Ga i family, except for
Ga t and Ga gust which simply do not have any N-terminal cysteines, palmitoylation