Purification and some properties of a protein factor binding and deacylating initiator transfer ribonucleic acid.
1. A protein factor promoting the binding of initiator tRNA to the 40S ribosomal subunit was purified to homogeneity (more than 2500-fold) from rat liver cytosol. It has a mol.wt. of 265000 and is composed of four subunits of identical molecular weight. 2. This factor directs the binding of methionyl-tRNA(fMet) and to a lesser extent also of N-acetylphenylalanyl-tRNA, but not of methionyl-tRNA(Met) or phenylalanyl-tRNA, to the smaller ribosomal subunit at high concentrations of GTP (8-10mm) with an optimum at pH4.0. As evidenced by sucrose-density-gradient centrifugation, initiator tRNA becomes bound to the 40S subunit or to 80S ribosomes. 3. A deacylase activity specific for methionyl-tRNA(fMet) is associated with the pure factor. The factor significantly stimulates the translation of natural message in systems containing polyribosomes and both purified peptide-elongation factors. 4. The factor binds initiator tRNA or GTP to form unstable binary complexes and forms a ternary complex with methionyl-tRNA(fMet) and GTP. This complex is relatively stable. 5. In the absence of any cofactors the factor forms a stable complex with 40S and 80S ribosomes. This preformed ribosomal complex binds efficiently initiator tRNA at pH7.5 and low concentrations of GTP (1-2mm). The ternary complex of the factor with methionyl-tRNA(fMet) and GTP may be liberated from this ribosomal complex. 6. A protein factor capable of promoting the binding and simultaneously the deacylation of initiator tRNA may apparently have a regulatory function in physiological gene translation by removing an excess of methionyl-tRNA(fMet) not required for translation.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel; D005260:Female; D006160:Guanosine Triphosphate / Q000378:metabolism; D008099:Liver / Q000032:analysis; D008297:Male; D008970:Molecular Weight; D010448:Peptide Initiation Factors / Q000302:isolation & purification / Q000378:metabolism; D011485:Protein Binding; D012343:RNA, Transfer / Q000378:metabolism*; D051381:Rats; D012270:Ribosomes / Q000378:metabolism
D010448:Peptide Initiation Factors; D006160:Guanosine Triphosphate; D012343:RNA, Transfer