Bimodal action of spermine on ribosomal peptidyltransferase at low concentration of magnesium ions.
At 6 mM Mg2+, submillimolar concentrations of spermine affect the end-point as well as the kinetic phase of puromycin reaction in a cell-free system from Escherichia coli. When the ternary complex AcPhe-tRNA-poly(U)-ribosome (complex C) is formed in the absence of ribosomal wash (FWR fraction), the final degree of AcPhe-puromycin synthesis is raised from 12% to 60%, as the concentration of spermine increases from zero to 200 microM. However, spermine displays partial noncompetitive inhibition at the kinetic phase of the reaction. The inhibitory effect of spermine is related with its binding to AcPhe-tRNA. When complex C is formed in the presence of FWR fraction, spermine slightly affects the final degree of puromycin synthesis is markedly stimulated by the addition of relatively low concentrations of spermine. Kinetic analysis of the activation phase revealed that spermine attached on a specific site of complex C, acts as a nonessential, partial noncompetitive activator. The stimulatory effect of spermine seems to be due to its interaction with ribosomes. Further additions of spermine cause partial noncompetitive inhibition on the puromycin reaction. This result suggests that complex C possesses a second binding site, responsible for the inhibitory effect of spermine. Both activator and inhibitor sites can be occupied by spermine at the same time.
Drainas
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Laboratory of Biochemistry, School of Medicine, University of Patras, Greece.
Kalpaxis
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D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D010458:Peptidyl Transferases / Q000378:metabolism*; D011072:Poly U / Q000378:metabolism; D011691:Puromycin / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012270:Ribosomes / Q000201:enzymology*; D013096:Spermine / Q000378:metabolism / Q000494:pharmacology*
D012346:RNA, Transfer, Amino Acyl; C006383:tRNA, N-acetylphenylalanine-; D011072:Poly U; D013096:Spermine; D011691:Puromycin; D010458:Peptidyl Transferases; D008274:Magnesium