Eukaryotic Translation

Translated Eukaryotic Genes in Which Introns Have Been Demonstrated... 

See also Initiation of Translation, Structure of tRNAs, Structure of Prokaryotic mRNAs, Termination of Translation, Eukaryotic vs Prokaryotic Translation (from Chapter 28)... 

RNA Structures. mRNAs contain the nucleotide sequence that is converted into the amino acid sequence of a protein in the process of translation. Eukaryotic mRNA has a structure known as a cap at the 5 -end, a sequence of adenine nucleotides (a poly(A) tail) at the 3 -end, and a coding region in the center containing codons that dictate the sequence of amino acids in a protein or relay a signal. Each codon in the genetic code is a different sequence of three nucleotides. 

Assemble two translation reactions from each concentration point, adding a well-translated eukaryotic mRNA to one. 

FIGURE 11.24 The properties of mRNA molecules in prokaryotic versus eukaryotic cells during transcription and translation. 

The primary transcripts generated by RNA polymerase II—one of three distinct nuclear DNA-depen-dent RNA polymerases in eukaryotes—are promptly capped by 7-methylguanosine triphosphate caps (Figure 35-10) that persist and eventually appear on the 5 end of mature cytoplasmic mRNA. These caps are necessary for the subsequent processing of the primary transcript to mRNA, for the translation of the mRNA, and for protection of the mRNA against exonucleolytic attack. 

Eukaryotic mRNA synthesis results in a pre-mRNA precursor that contains extensive amounts of excess RNA (introns) that must be precisely removed by RNA splicing to generate functional, translatable mRNA composed of exonic coding and noncoding sequences. 

Initiation of protein synthesis requires that an mRNA molecule be selected for translation by a ribosome. Once the mRNA binds to the ribosome, the latter finds the correct reading frame on the mRNA, and translation begins. This process involves tRNA, rRNA, mRNA, and at least ten eukaryotic initiation factors (elFs), some of which have multiple (three to eight) subunits. Also involved are GTP, ATP, and amino acids. Initiation can be divided into four steps (1) dissociation of the ribosome into its 40S and 60S subunits (2) binding of a ternary complex consisting of met-tRNAf GTP, and eIF-2 to the 40S ribosome to form a preinitiation complex (3) binding of mRNA to the 40S preinitiation complex to form a 43S initiation complex and (4) combination of the 43S initiation complex with the 60S ribosomal subunit to form the SOS initiation complex. 

Kozak M Structural features in eukaryotic mRNAs that modulate the initiation of translation. J Biol Chem 1991 266 1986. 

Sachs AB, Sarnow P, Hentze MW Starting at the beginning, middle and end translation Initiation in eukaryotes. Cell 1997 98 831. 

Previously, it has been reported that the amounts of eukaryotic initiation factors in wheat germ extract prepared by a common method were deficient for the translation of some kinds of mRNAs including a-amylase mRNA and (i-globin mRNA [2]. Therefore, it can be expected that the activity of wheat germ extract prepared by a common method can be enhanced by the simple addition of extract containing deficient initiation factors. In this study, a wheat germ extract was further purified partially by ammonium sulfate fractionation... 

The catalytic activities of the fortified wheat germ cell-free systems supplemented with each fraction were investigated (Fig. 2). As shown in Fig. 2, only 0 - 40 % ammonium sulfate fraction showed an enhancement in DHFR protein synthesis. This enhancement of protein experimental results and the fact that the various eukaryotic initiation factors are contained in synthesis was also confirmed by SDS-PAGE and autoradiography (Fig. 3). From the above 0-40 % ammonium sulfate fraction [5, 6], it can be concluded that the amount of initiation factors in a conventionally prepared wheat germ cell-fi extract is deficient for the translation of DHFR with internal ribosome entry site. Therefore, it needs to supplement a wheat germ cell-free extract with the fraction containing the limited initiation factors for the efficient protein translation, and this fortified cell-free system can be easily made by simple... 

Post-translational modification of proteins plays a critical role in cellular function. For, example protein phosphorylation events control the majority of the signal transduction pathways in eukaryotic cells. Therefore, an important goal of proteomics is the identification of post-translational modifications. Proteins can undergo a wide range of post-translational modifications such as phosphorylation, glycosylation, sulphonation, palmitoylation and ADP-ribosylation. These modifications can play an essential role in the function of the protein and mass spectrometry has been used to characterize such modifications. 

The mDHFR protein complementation assay has been used to map a signal transduction network that controls the initiation of translation in eukaryotes (Remy and Michnick, 2001). A total of 35 different pairs of full-length proteins were analyzed and 14 interactions were identified using the survival selection of cells grown in the absence of nucleotides. In addition, the use of the fMTX reagent in combination with fluorescence microscopy was used to localize the protein complex within cells (Remy and Michnick, 2001). 

In contrast to the photosynthetic eukaryotes, photoprotection in cyanobacteria is not induced by the presence of a transthylakoid ApH or the excitation pressure on PSII. Instead, intense blue-green light (400-550 nm) induces a quenching of PSII fluorescence that is reversible in minutes even in the presence of translation inhibitors (El Bissati et al. 2000). Fluorescence spectra measurements and the study of the NPQ mechanism in phycobilisome- and PSII-mutants of the cyanobacterium Synechocystis PCC6803 indicate that this mechanism involves a specific decrease of the fluorescence emission of the phycobilisomes and a decrease of the energy transfer from the phycobilisomes to the RCs (Scott et al. 2006, Wilson et al. 2006). The site of the quenching appears to be the core of the phycobilisome (Scott et al. 2006, Wilson et al. 2006, Rakhimberdieva et al. 2007b). 

Figure 7.4 (a) IREs in eukaryotic mRNAs the secondary structures of ferritin and transferrin receptor IREs. (b) The IRE localization in mRNAs the translation/ribosome binding element in the 5 -UTR of ferritin mRNA is above, that of the stability/ turnover element in the 3 -UTR of transferrin receptor mRNA is below. Adapted from Theil, 1998, by courtesy of Marcel Dekker, Inc. 

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