Protein initiation factors

Figure 1.8 Translation of messenger RNA. The attachment of a ribosome to the mRNA involves protein initiation factors and the recognition of a particular base sequence, the start codon. A single mRNA can be simultaneously translated by more than one ribosome, forming a polyribosome. Synthesis occurs in the direction from the 5 end of messenger RNA to the 3 end. For further details of protein synthesis see Chapter 20.

The first phase of translation, initiation, involves several steps. First, two proteins, initiation factors IF-1 and IF-3, bind to the 30 S subunit (1). Another factor, IF-2, binds as a complex with GTP (2). This allows the subunit to associate with the mRNA and makes it possible for a special tRNA to bind to the start codon (3). In prokaryotes, this starter tRNA carries the substituted amino acid N-formylmethionine (fMet). In eukaryotes, it carries an unsubstituted methionine. Finally, the 50 S subunit binds to the above complex (4). During steps 3 and 4, the initiation factors are released again, and the GTP bound to IF-2 is hydrolyzed to GDP and Pj. 

Increasing evidence suggests that Met-tRNAf is also the initiator tRNA in eukaryotic systems (C35, G16). The failure of previous experiments to demonstrate the role of this Met-tRNAf for the in vitro protein synthesis is probably due to the lack of protein initiation factors. Mi, Ma, and Ms, which are present in a ribosomal salt-wash protein fraction (P24, S35, S36). The most recent experiments by Anderson and co-workers (C34, C35) show that the Met-tRNAf binds the initiation factors Ml and Ms to form an initiation complex with messenger RNA. The binding of this complex requires CTP and Mg + ions. A methionyl-valine dipeptide production is the next step in the biosynthesis of the chain the synthesis of this bond requires Mg + ions, an additional initiation... 

Initiation. Protein synthesis in bacteria begins by the association of one 308 subunit (not the 708 ribosome), an mRNA, a charged tRNA , three protein initiation factors, and guanosine 5 -triphosphate (GTP). These molecules make up the 308 preinitiation complex. Association occurs at an initiator AUG codon, whose selection was described above. A 508 subunit joins to the 308 subunit to form a 708 initiation complex (Figure 25-11). This joining process requires hydrolysis of the GTP contained in the 308 preinitiation complex. There are two tRNA... 

Let us look more cl osely at the molecular events at the ribosome, where the recognition and formation of peptide bonds actually occurs. We have seen that the ribosome is composed of two different subunits, but only one of these subunits, the smaller of the two, is essential for initiation of protein synthesis, although it must be associated with the larger unit before chain elongation can proceed. Initiation also requires the presence of an energy source (supplied not by ATP but GTP), a particular amino acyl t-RNA whose anti-codon corresponds to the start here codon on m-RNA and, at least in bacteria, three soluble protein initiation factors called IF1, IF2, IF3. The ribosome has two sites for t-RNA binding, the P site and the A site, but only initiator t-RNA can bind to the P site - all other incoming amino acyl t-RNAs bind to the A site. 

The major differences between prokaryotic and eukaryotic translation control mechanisms are related to the complexity of eukaryotic gene expression. Features that distinguish eukaryotic translation include mRNA export (spatial separation of transcription and translation), mRNA stability (the half-lives of mRNA can be modulated), negative translational control (the translation of certain mRNAs can be blocked by the binding of specific repressor proteins), initiation factor phosphorylation (mRNA translation rates are altered by certain circumstances when eIF-2 is phosphorylated), and translational frame-shifting (certain mRNAs can be frame-shifted so that a different polypeptide is synthesized). 

The initiation complex in E. coli requires mRNA, the SOS ribosomal subunit, fmet-tRNA S GTP, and three protein-initiation factors, called IF-1, IF-2, and IF-3. The IF-3 protein is needed for the binding of mRNA to the ribosomal subunit. The other two protein factors are required for the binding of fmet-tRN fmet mRNA-30S complex. 

Fig. 2. Formation of a stable initiation complex between a 70 S ribosome and messenger RNA. In the final complex fMet-tRNAf " is in the correct position for the formation of a peptide bond. IF-1, IF-2, and IF-3 are the protein initiation factors and fMet-tRNAf " is the formyl methionyl tRNA which is used for the initiation of protein synthesis in prokaryotes. The process in animal cells is thought to be substantially the same, the initiation factors being termed IF-Ml, IF-M2, and IF-M3 and the initiator tRNA, Met-tRNAt . The methionine attached to this tRNA species is not normally formylated but can be so modified by enzymes from bacterial cells.

Protein initiation factors Eukaryotic initiation factors (elFs) Initiation factors (IPs)... 

Thus, the initiation process in bacteria involves the recognition of a special codon by the anticodon of an fMet-tRNA. Yet a number of other factors are needed for initiation, including at least three protein initiator factors—Fi (mol wt 9,000), F2 (mol wt 65,000), F3 (mol wt 24,000)—and GTP. 

Protein-initiating factors which were extracted from unwashed ribosomes promoted the formation of SOS complexes only when the mRNA used was extracted from individuals in the same developmental stage. The specificity of protein-initiating factors determines the control mechanism of protein synthesis by ribosomal selection of mRNA for translation. 

Initiation of protein synthesis requires assembly of the ribosomal subunits, messenger RNA, and initiator tRNA at the start codon. This organization of translational components is facilitated by protein initiation factors (Fig. 5). 

A modifier-specifier role for proteins, wherein a new biologic function or enhanced activity is achieved, may well be more common than is presently appreciated. For example, factors V and VIII in the blood coagulation process appear to act in this way (Davie et al., 1979), as do specific protein initiation factors which participate in protein synthesis (Weissbach and Ochoa, 1976). 

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