Harpers

PROTEIN SYNTHESIS AND THE GENETIC CODE / 367E. FORMATION OF THE 80S INITIATION COMPLEXThe binding of the 60S ribosomal subunit to the 48Sinitiation complex involves hydrolysis of the GTPbound to eIF-2 by eIF-5. This reaction results in releaseof the initiation factors bound to the 48S initiationcomplex (these factors then are recycled) and the rapidassociation of the 40S and 60S subunits to form the80S ribosome. At this point, the met-tRNA i is on the Psite of the ribosome, ready for the elongation cycle tocommence.The Regulation of eIF-4E Controlsthe Rate of InitiationThe 4F complex is particularly important in controllingthe rate of protein translation. As described above, 4F isa complex consisting of 4E, which binds to the m 7 Gcap structure at the 5′ end of the mRNA, and 4G,which serves as a scaffolding protein. In addition tobinding 4E, 4G binds to eIF-3, which links the complexto the 40S ribosomal subunit. It also binds 4A and4B, the ATPase-helicase complex that helps unwind theRNA (Figure 38–7).4E is responsible for recognition of the mRNA capstructure, which is a rate-limiting step in translation.This process is regulated at two levels. Insulin and mitogenicgrowth factors result in the phosphorylation of4E on ser 209 (or thr 210). Phosphorylated 4E binds tothe cap much more avidly than does the nonphosphorylatedform, thus enhancing the rate of initiation. Acomponent of the MAP kinase pathway (see Figure43–8) appears to be involved in this phosphorylationreaction.The activity of 4E is regulated in a second way, andthis also involves phosphorylation. A recently discoveredset of proteins bind to and inactivate 4E. Theseproteins include 4E-BP1 (BP1, also known as PHAS-1)and the closely related proteins 4E-BP2 and 4E-BP3.BP1 binds with high affinity to 4E. The [4E]•[BP1] associationprevents 4E from binding to 4G (to form 4F).Since this interaction is essential for the binding of 4Fto the ribosomal 40S subunit and for correctly positioningthis on the capped mRNA, BP-1 effectively inhibitstranslation initiation.Insulin and other growth factors result in the phosphorylationof BP-1 at five unique sites. Phosphorylationof BP-1 results in its dissociation from 4E, and itcannot rebind until critical sites are dephosphorylated.The protein kinase responsible has not been identified,but it appears to be different from the one that phosphorylates4E. A kinase in the mammalian target ofrapamycin (mTOR) pathway, perhaps mTOR itself, isinvolved. These effects on the activation of 4E explainin part how insulin causes a marked posttranscriptional4E-BP 4E-BPeIF-4E4FCapInsulin(kinaseactivation)PO 4AUGeIF-4EPO 4eIF-4GeIF-4AeIF-4G eIF-4FcomplexeIF-4EeIF-4APO 4(A) nFigure 38–7. Activation of eIF-4E by insulin and formationof the cap binding eIF-4F complex. The 4F-capmRNA complex is depicted as in Figure 38–6. The 4Fcomplex consists of eIF-4E (4E), eIF-4A, and eIF-4G. 4E isinactive when bound by one of a family of binding proteins(4E-BPs). Insulin and mitogenic factors (eg, IGF-1,PDGF, interleukin-2, and angiotensin II) activate a serineprotein kinase in the mTOR pathway, and this results inthe phosphorylation of 4E-BP. Phosphorylated 4E-BPdissociates from 4E, and the latter is then able to formthe 4F complex and bind to the mRNA cap. Thesegrowth peptides also phosphorylate 4E itself by activatinga component of the MAP kinase pathway. Phosphorylated4E binds much more avidly to the cap thandoes nonphosphorylated 4E.increase of protein synthesis in liver, adipose tissue, andmuscle.Elongation Also Is a Multistep Process(Figure 38–8)Elongation is a cyclic process on the ribosome in whichone amino acid at a time is added to the nascent peptidechain. The peptide sequence is determined by the orderof the codons in the mRNA. Elongation involves severalsteps catalyzed by proteins called elongation factors (EFs).These steps are (1) binding of aminoacyl-tRNA to the Asite, (2) peptide bond formation, and (3) translocation.