TY - JOUR
T1 - The RNA-binding properties of protein synthesis initiation factor eIF-2
AU - Flynn, Andrea
AU - Shatsky, Ivan N.
AU - Proud, Christopher G.
AU - Kaminski, Ann
N1 - Funding Information:
This work was supported by Project Grant (to C.G.P.) from the Medical Research Council. We are grateful to Drs. Sue Oldfield and Gavin Welsh (Bristol) for preparing the elF-2 and elF-2B used in these experiments. We are also grateful to Sue Oldfield and Richard Jackson (Cambridge) for helpful discussions and valuable technical advice. The joint experiments with Dr. Shatsky were made possible by a European Interlaboratory Collaboration Grant from the Wellcome Trust, who supported his visit to Bristol.
PY - 1994/10/18
Y1 - 1994/10/18
N2 - Protein synthesis initiation factor eIF-2 bound ATP in the presence or absence of Mg2+ ions. ATP impaired the binding of GTP or GDP to eIF-2. However, excess GTP did not significantly decrease the binding of ATP to eIF-2, suggesting eIF-2 has distinct ATP and GTP binding sites. Highly purified eIF-2 can bind mRNA, and this did not require the mRNA to be capped. mRNA binding was saturable, and maximal binding corresponded to about 0.4 mol mRNA bound per mol eIF-2. GTP, and, at lower concentrations, GDP, inhibited the binding of mRNA to eIF-2. In addition, ATP and other nucleoside triphosphates decreased mRNA binding. The implications of these findings for the structure and function of eIF-2 are discussed. Preparations of eIF-2 deficient in the β-subunit showed reduced ability to bind mRNA, suggesting that while it is not essential for mRNA binding, this subunit is involved in the interaction. Consistent with this is the observation that ultraviolet crosslinking of mRNA to eIF-2 resulted primarily in labelling of the β-subunit. Subsequent analysis revealed that mRNA was cross-linked to the C-terminal region of eIF-2b which contains a putative Zn-finger structure.
AB - Protein synthesis initiation factor eIF-2 bound ATP in the presence or absence of Mg2+ ions. ATP impaired the binding of GTP or GDP to eIF-2. However, excess GTP did not significantly decrease the binding of ATP to eIF-2, suggesting eIF-2 has distinct ATP and GTP binding sites. Highly purified eIF-2 can bind mRNA, and this did not require the mRNA to be capped. mRNA binding was saturable, and maximal binding corresponded to about 0.4 mol mRNA bound per mol eIF-2. GTP, and, at lower concentrations, GDP, inhibited the binding of mRNA to eIF-2. In addition, ATP and other nucleoside triphosphates decreased mRNA binding. The implications of these findings for the structure and function of eIF-2 are discussed. Preparations of eIF-2 deficient in the β-subunit showed reduced ability to bind mRNA, suggesting that while it is not essential for mRNA binding, this subunit is involved in the interaction. Consistent with this is the observation that ultraviolet crosslinking of mRNA to eIF-2 resulted primarily in labelling of the β-subunit. Subsequent analysis revealed that mRNA was cross-linked to the C-terminal region of eIF-2b which contains a putative Zn-finger structure.
KW - Cross-linking
KW - RNA binding
KW - Zinc finger
KW - eIF-2
UR - http://www.scopus.com/inward/record.url?scp=0028110186&partnerID=8YFLogxK
U2 - 10.1016/0167-4781(94)90051-5
DO - 10.1016/0167-4781(94)90051-5
M3 - Article
C2 - 7918624
AN - SCOPUS:0028110186
SN - 0167-4781
VL - 1219
SP - 293
EP - 301
JO - BBA - Gene Structure and Expression
JF - BBA - Gene Structure and Expression
IS - 2
ER -