The role of cholesteryl 14‐methylhexadecanoate in the function of eukaryotic peptide elongation factor 1

Abstract

The binding of [3H]cholesteryl 14-methylhexadecanoate by a highly purified peptide elongation factor 1 from rabbit reticulocytes is significantly enhanced by GTP and CTP, much less by guanosine S-[P,y-methylene]-triphosphate and not at all by ATP or UTP. Removal of endogenous cholesteryl14-methylhexadecanoate present in the molecule of the factor [Hradec, J. et al. (1971) Biochem. J. 123, 959-9661 by digestion with immobilized cholesterol esterase resulted in an almost complete loss of GTPase activity and this could be restored to nearly normal values by the addition of the ester. The same holds true for the GTP-dependent autophosphorylation of the protein-synthesis factor. Cholesteryl 14-methylhexadecanoate was bound only by the fl subunit of the factor. Addition of the CI subunit, which was inactive on its own, stimulated the binding of the ester to the fl subunit in a sigmoid dependance. The binding of the ester was significantly stimulated by aminoacyl-tRNA but this effect was fully abolished by sodium fluoride, indicating a relation of cholesteryl 14-methylhexadecanoate to the dephosphorylation of the peptide elongation factor. Treatment of the factor with cholesterol esterase decreased its activity in the poly(U)-dependent binding of phenylalanyl-tRNA to ribosome and this activity was again restored by the addition of cholesteryl 14-methylhexadecanoate. The ester thus interacts with the GTP-dependent autophosphorylation of peptide elongation factor 1 and in this way modulates the activity of the factor. A putative scheme is presented explaining the mode of action of cholesteryl 14-methylhexadecanoate. Cholesteryl 14-methyldecanoate (CMH) was found as an essential component in the molecule of several enzymes and protein factors involved in eukaryotic protein synthesis. Its presence modulates the activity of these preparations and is required for their normal function (see [l] for a review).Extraction of partially purified eEF-1 with organic solvents [2] or its treatment with cholesterol esterase [3] resulted in a decreased binding of aminoacyl-tRNA to ribosomes in the presence of such factor preparations and their activity was completely restored by the addition of CMH. Labeled CMH became bound to eEF-1 and exhibited specific allosteric interactions between added exogenous ester molecules and those endogenously present in peptide initiation and peptide elongation factors [4]. These interactions apparently represented homotropic interactions between the molecules of CMH originally present in the factor and molecules of exogenous ester that became bound by the factor. Thus CMH may be considered as a ligand modulating the activity of protein-synthesis factors by allosteric effects [5].Preliminary experiments on the mode of action of CMH in protein synthesis indicated that this ester is able to interfere with the phosphorylation of protein-synthesis factors (TuhaEkova, Z., unpublished). The GTP-dependent autophosphorylation of peptide-initiation [6] and peptide-elongation factors [7] has been demonstrated as ...

Monomethyl Branched-Chain Fatty Acids Play an Essential Role in Caenorhabditis elegans Development

“…A removal of this FA from its protein counterparts results in a loss of hydrophobicity (Jones and Rivett 1997). Other studies indicated that C17anteISO esterified to cholesterol binds to and activates enzymes of protein biosynthesis (Tuhackova and Hradec 1985; Hradec and Dufek 1994). A potential significance of mmBCFAs for human health is associated with a long-observed correlation between amounts of these FAs and disease conditions such as brain deficiency (Ramsey et al 1977) and cancer (Hradec and Dufek 1994).…”

Translational dynamics

“…The mechanism responsible for the dissociation is not unterstood at present but guanosine nucleotides and treament with protease or phospholipase lead to a disaggregation [123,1301. eEF-1/3 seems to be responsible for the formation of eEF-1, and the function of eEF-lP is dependent on a cholesteryl 14-methylhexadecanoate [131]. The recent observation that eEF-la contains ethanolamine and glycerol may indicate that the protein is anchored to membranes or to the cytoskeleton [132,133].…”

“…suggesting that the interface is the 'active site' of the ribosome. [12,131 with the modification that the position of the platform is assumed to be equivalent to that in the prokaryotic small ribosomal subunit [4] Also neutron-scattering experiments and the electron distribution indicate a peripheral localization of the proteins with an rRNA core at the centre of the ribosomal particle [23-281. Thus, the interface between the two ribosomal particles contains few proteins [29] and a direct contact between the two major ribosomal RNAs across the subunit interface has been observed [30].…”