Asparagine-135 of elongation factor Tu is a crucial residue for the folding of the guanine nucleotide binding pocket.
This work studies the structure-function relationships of Asn135, a residue situated in the GTP binding pocket of elongation factor Tu (EF-Tu). For this purpose we constructed EF-TuN135D/D138N and assayed its reactivity towards various purine nucleotides. We found that EF-TuN135D/D138N had no functional effect with GTP, ATP, XTP and isoGTP. The lack of a productive interaction with isoGTP shows that the Asn135 side-chain does not recognize the exocyclic keto group of the guanine base. However, EF-TuN135D/D138N, whose native conformation is stabilized by either elongation factor Ts or kirromycin, was able to support the enzymatic binding of aa-tRNA to the ribosome in the absence of any nucleotide, when in complex with the antibiotic. Taken together, these results show that Asn135 is important for the correct folding of the nucleotide binding site and that EF-Tu.kirromycin can mediate the binding of aa-tRNA to the mRNA-programmed ribosomes independently of the native conformation of this site.
SDI n. 61840 du CNRS, Laboratoire de Biochimie, Ecole Polytechnique, Palaiseau, France.
A
A
Weijland
R
R
Sarfati
O
O
Bârzu
A
A
Parmeggiani
D001216:Asparagine / Q000737:chemistry*; D001483:Base Sequence; D001665:Binding Sites; D006160:Guanosine Triphosphate / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides; D010444:Peptide Elongation Factor Tu / Q000737:chemistry*; D017510:Protein Folding; D013329:Structure-Activity Relationship
D009838:Oligodeoxyribonucleotides; D001216:Asparagine; D006160:Guanosine Triphosphate; D010444:Peptide Elongation Factor Tu