title: Testing with puromycin and amino acyl tRNAs that limit the rate of peptide chain extension.
abstract: With puromycin one can recognize when the synthesis of a given protein is dependent on amino acyl tRNA that is present in rate limiting amount. We demonstrate this use of puromycin by its interaction with another inhibitor, L-o-methylthreonine. L-o-methylthreonine lowers the Ile-tRNA concentration in the cell, thereby inhibiting synthesis of proteins containing isoleucine. In certain rabbits, the alpha hemoglobin chain has three isoleucyl residues and the beta chain none. L-o-methylthreonine thus inhibits alpha globin synthesis in intact reticulocytes from these rabbits. When puromycin and L-o-methylthreonine are used together, the two inhibitors synergize in inhibiting alpha globin synthesis. Hence, puromycin is a more effective inhibitor when the Ile-tRNA concentration is lowered. Cycloheximide and sodium fluoride have different modes of action from puromycin. Neither synergizes with L-o-methylthreonine; instead, the interaction is less than additive. We have found that beta chain synthesis in rabbit reticulocytes is more sensitive than alpha to inhibition by puromycin. This difference could reflect either differences in amino acid sequence or tRNA dependent limitations of beta chain elongation. The switch from fetal to adult hemoglobin in humans does not involve changes in limiting amino acyl tRNA because, for cord blood from infants of different developmental ages, the puromycin sensitivity of incorporation into gamma and beta chains remains constant.
mesh_terms: D000818:Animals; D003513:Cycloheximide / Q000494:pharmacology; D004357:Drug Synergism; D005319:Fetal Hemoglobin / Q000096:biosynthesis*; D005459:Fluorides / Q000494:pharmacology; D005914:Globins / Q000096:biosynthesis*; D006801:Humans; D007532:Isoleucine / Q000378:metabolism; D046911:Macromolecular Substances; D010441:Peptide Chain Elongation, Translational / Q000187:drug effects*; D011691:Puromycin / Q000494:pharmacology*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism*; D011817:Rabbits; D013912:Threonine / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D046911:Macromolecular Substances; D012346:RNA, Transfer, Amino Acyl; D007532:Isoleucine; D013912:Threonine; D011691:Puromycin; D005914:Globins; D005319:Fetal Hemoglobin; D003513:Cycloheximide; D005459:Fluorides
references.citation: J Theor Biol. 1974 Jan;43(1):211-25
references.pmid: 4591738
references.citation: Arch Biochem Biophys. 1968 Mar 20;124(1):306-13
references.pmid: 4298495
references.citation: J Mol Biol. 1965 Feb;11:358-72
references.pmid: 14290351
references.citation: Proc Natl Acad Sci U S A. 1964 Jun;51:1134-41
references.pmid: 14215634
references.citation: Proc Natl Acad Sci U S A. 1964 Apr;51:585-92
references.pmid: 14166766
references.citation: J Mol Biol. 1969 Jul 14;43(1):123-33
references.pmid: 5811817
references.citation: Biochim Biophys Acta. 1974 Jun 7;351(2):437-41
references.pmid: 4407315
references.citation: Proc Natl Acad Sci U S A. 1959 Dec;45(12):1721-9
references.pmid: 16590564
references.citation: Proc Natl Acad Sci U S A. 1961 Mar 15;47:247-61
references.pmid: 13723017
references.citation: Nature. 1976 Mar 4;260(5546):67-70
references.pmid: 1264198
references.citation: J Mol Biol. 1966 Oct 28;21(1):51-69
references.pmid: 5969765
references.citation: J Biol Chem. 1971 Jan 10;246(1):174-81
references.pmid: 5541758
references.citation: J Mol Biol. 1968 Aug 28;36(1):31-45
references.pmid: 5760537
references.citation: Genetics. 1974 Jan;76(1):99-108
references.pmid: 4818267
references.citation: Blut. 1958 Nov;4(5):241-9
references.pmid: 13607582
references.citation: J Biol Chem. 1974 Jul 25;249(14):4594-600
references.pmid: 4843145
references.citation: Ann N Y Acad Sci. 1974 Nov 29;241(0):223-31
references.pmid: 4530654
references.citation: Jinrui Idengaku Zasshi. 1967 Oct;12(2):55-61
references.pmid: 4866959
references.citation: J Mol Biol. 1966 Aug;19(1):91-108
references.pmid: 5967288
references.citation: Eur J Biochem. 1970 Sep;16(1):136-42
references.pmid: 4917228
references.citation: Arch Biochem Biophys. 1973 Oct;158(2):922-4
references.pmid: 4782539
references.citation: Biochim Biophys Acta. 1976 Dec 1;454(2):329-37
references.pmid: 1033768
references.citation: Biochemistry. 1967 Jun;6(6):1704-11
references.pmid: 5340946
references.citation: Eur J Biochem. 1975 Oct 15;58(2):339-50
references.pmid: 1183442
references.citation: J Mol Biol. 1968 Apr 14;33(1):315-8
references.pmid: 5646650
references.citation: Proc Natl Acad Sci U S A. 1975 Jan;72(1):258-62
references.pmid: 1054502
references.citation: Proc Natl Acad Sci U S A. 1969 Aug;63(4):1206-13
references.pmid: 5260921
references.citation: Proc Natl Acad Sci U S A. 1968 Apr;59(4):1349-55
references.pmid: 5240035
references.citation: J Biol Chem. 1972 Jun 10;247(11):3622-9
references.pmid: 4624122
references.citation: Hoppe Seylers Z Physiol Chem. 1966;347(4):207-11
references.pmid: 5991669
references.citation: Hoppe Seylers Z Physiol Chem. 1968 Aug;349(8):1073-5
references.pmid: 5679924