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Mutational analysis of conserved positions potentially important for initiator tRNA function in Saccharomyces cerevisiae
Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
1992 (Engelska)Ingår i: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 12, nr 4, s. 1432-1442Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The conserved positions of the eukaryotic cytoplasmic initiator tRNA have been suggested to be important for the initiation of protein synthesis. However, the role of these positions is not known. We describe in this report a functional analysis of the yeast initiator methionine tRNA (tRNA(iMet)), using a novel in vivo assay system which is not dependent on suppressor tRNAs. Strains of Saccharomyces cerevisiae with null alleles of the four initiator methionine tRNA (IMT) genes were constructed. Consequently, growth of these strains was dependent on tRNA(iMet) encoded from a plasmid-derived gene. We used these strains to investigate the significance of the conserved nucleosides of yeast tRNA(iMet) in vivo. Nucleotide substitutions corresponding to the nucleosides of the yeast elongator methionine tRNA (tRNA(MMet)) have been made at all conserved positions to identify the positions that are important for tRNA(iMet) to function in the initiation process. Surprisingly, nucleoside changes in base pairs 3-70, 12-23, 31-39, and 29-41, as well as expanding loop I by inserting an A at position 17 (A17) had no effect on the tester strain. Nucleotide substitutions in positions 54 and 60 to cytidines and guanosines (C54, G54, C60, and G60) did not prevent cell growth. In contrast, the double mutation U/rT54C60 blocked cell growth, and changing the A-U base pair 1-72 to a G-C base pair was deleterious to the cell, although these tRNAs were synthesized and accepted methionine in vitro. From our data, we suggest that an A-U base pair in position 1-72 is important for tRNA(iMet) function, that the hypothetical requirement for adenosines at positions 54 and 60 is invalid, and that a U/rT at position 54 is an antideterminant distinguishing an elongator from an initiator tRNA in the initiation of translation.

Ort, förlag, år, upplaga, sidor
American Society for Microbiology , 1992. Vol. 12, nr 4, s. 1432-1442
Nationell ämneskategori
Mikrobiologi inom det medicinska området
Identifikatorer
URN: urn:nbn:se:umu:diva-32176DOI: 10.1128/MCB.12.4.1432ISI: A1992HK75100004PubMedID: 1549105OAI: oai:DiVA.org:umu-32176DiVA, id: diva2:301474
Tillgänglig från: 2010-03-03 Skapad: 2010-03-03 Senast uppdaterad: 2019-01-23Bibliografiskt granskad

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von Pawel-Rammingen, UlrichByström, Anders S

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