Eukaryotic initiation factor 4A is the component that interacts with ATP in protein chain initiation.

AUTOR(ES)

Seal, S N

RESUMO

Protein synthesis in a resolved homogenate of wheat germ requires ATP and eight factors functioning at the level of protein chain initiation. To identify the component(s) interacting with ATP, the different factors were treated with the ATP affinity analogue 5'-p-fluorosulfonylbenzoyladenosine (FSBA) and tested for their function in protein synthesis. The activity of eukaryotic initiation factor 4A (eIF4A) was strongly curtailed, whereas all other factors were unaffected. At a concentration of 250 microM, AMP, ADP, and ATP protected eIF4A against FSBA inactivation, whereas at a concentration 50 microM, protection was afforded only by ATP. GTP did not protect at a concentration of 250 microM. In another approach, the substrate analogue 2',3'-O-(2,4,6-trinitrophenyl)adenosine 5'-triphosphate (TNP-ATP) was found to inhibit protein synthesis in a manner, at least in part, competitive with ATP. Supplementing a TNP-ATP inhibited reaction with eIF4A substantially reversed the inhibition. Except for a small effect by factor C1, no reversal was obtained with any other component. Finally, a preincubation of ribosomes with ATP, mRNA, and eIF4A resulted in the formation of a complex capable of TNP-ATP-resistant amino acid incorporation. These data are interpreted to indicate that the primary interaction of ATP is with eIF4A. A model is proposed reconciling this conclusion with other observations relevant to the mRNA . ribosome attachment reaction.

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