Effect of the nucleotide-37 on the interaction of tRNA(Phe) with the P site of Escherichia coli ribosomes.
The method of anticodon loop replacement has been used to make derivatives of yeast tRNA(Phe) with the substitution at position 37 (tRNA(Phe)GAAA) and at the anticodon(tRNA(Phe)GCAG). A quantitative study of the interaction of various types of deacylated yeast tRNA(Phe) (tRNA(Phe)+Y, tRNA(Phe)GAAA, tRNA(Phe)-Y) with the P site of the [70S ribosome*poly(U)]-complex was carried out at different Mg2+ concentrations and temperatures. The presence and nature of the nucleotide situated at the 3'-end of the anticodon are essential for such interaction in E coli ribosomes. Replacement of the Y base with the unmodified adenosine decreases the interaction enthalpy from 39 kcal/mol to 24 kcal/mol, whereas its removal reduces the interaction enthalpy to 16 kcal/mol. Replacement of the second anticodon nucleotide, adenosine, with cytosine further reduces the enthalpy to 6 kcal/mol, which is typical of tRNA-P site interaction in the absence of poly(U). In the absence of poly(U) the affinity of tRNA(PheY) for the P site of the 70S ribosome is five times lower than the affinity of tRNA(Phe+Y) or tRNA(Phe)GCAG. Thus, in the ribosome the modified nucleotide stabilizes the codon-anticodon interaction through its stacking interaction with the codon-anticodon base stack. In addition, this decreases the free energy of binding as a result of the interaction of the modified nucleotide itself with the hydrophobic center of the P site.
Katunin
V
V
Petersburg Nuclear Physics Institute, Russian Academy of Science, Leningrad Region.
Soboleva
N
N
Mahkno
V
V
Sedelnikova
E
E
Zhenodarova
S
S
Kirillov
S
S
D000215:Acylation; D000926:Anticodon; D001483:Base Sequence; D001665:Binding Sites; D003062:Codon; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism*; D008969:Molecular Sequence Data; D012329:RNA, Bacterial / Q000737:chemistry / Q000378:metabolism; D012360:RNA, Transfer, Phe / Q000737:chemistry / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D000926:Anticodon; D003062:Codon; D012329:RNA, Bacterial; D012360:RNA, Transfer, Phe