Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Study on the Function of Translation Initiation Factor IF1
Stockholm University, Faculty of Science, Department of Genetics, Microbiology and Toxicology.
2006 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Initiation is the first step in protein biosynthesis representing a fundamental event in cell life which determines fidelity, efficiency and regulation of gene expression. In addition to the ribosome and mRNA, three protein factors IF1, IF2 and IF3 are involved in the initiation of translation in prokaryotes. Several minor functions have been attributed to the smallest of these factors, IF1. However, the main function of IF1 remains to be elucidated.

In order to investigate the role of this protein in the initiation process we have mutated the corresponding gene infA. Using a high-copy plasmid and site-directed mutagenesis, the six arginine residues of IF1 were separately altered to leucine or aspartate. Another set of plasmid-encoded IF1 mutants with a cold-sensitive phenotype was collected using localized random mutagenesis. This strategy was followed by deletion of the chromosomal infA gene. All variants with a mutated infA gene on a plasmid and a deletion of the chromosomal infA copy were viable, except for an R65D alteration. Several of the mutated infA genes were successfully recombined into the chromosome thereby replacing the wild-type allele. Some of these mutants displayed reduced growth rates and a partial cold-sensitive phenotype.

The influence of the leucine group of mutants in IF1 on the expression of two reporter genes with different initiation and/or +2 codons has been investigated. Our results do not indicate any involvement of IF1 in recognition of the +2 codon immediately following the start codon, thus representing the A-site. In addition, this group of mutants has no changed efficiency of decoding at the near-cognate initiation codons UUG and GUG. However, one cold-sensitive IF1 mutant shows a general overexpression of both reporter genes, in particular at low temperatures. Overall, the results do not support the hypothesis that IF1 could possess codon discriminatory functions while blocking the A-site of the ribosome.

In this study we also identify that IF1 has RNA chaperone activity both in vitro and in vivo. The chaperone assays are based on splicing of the group I intron in the thymidylate synthase gene (td) from phage T4. Some of the IF1 mutant variants are more active as RNA chaperones than the wild-type. Both wild-type IF1 and mutant variants bind with high affinity to RNA in a band-shift assay. It is suggested that the RNA chaperone activity of IF1 contributes to RNA rearrangements during the early phase of translation initiation.

Place, publisher, year, edition, pages
Stockholm: Institutionen för genetik, mikrobiologi och toxikologi , 2006. , 65 p.
Keyword [en]
translation, initiation factor IF1, mutagenesis, A-site, RNA chaperone
National Category
Natural Sciences
Identifiers
URN: urn:nbn:se:su:diva-1032ISBN: 91-7155-216-2 (print)OAI: oai:DiVA.org:su-1032DiVA: diva2:189268
Public defence
2006-06-01, Nordenskiöldsalen, Geovetenskapens hus, Svante Arrhenius väg 8 C, Stockholm, 13:00
Opponent
Supervisors
Available from: 2006-05-08 Created: 2006-05-08Bibliographically approved
List of papers
1. Generation and characterization of functional mutants in the translation initiation factor IF1 of Escherichia coli
Open this publication in new window or tab >>Generation and characterization of functional mutants in the translation initiation factor IF1 of Escherichia coli
Show others...
2004 In: European Journal of Biochemistry, ISSN 1742-4658, Vol. 271, no 3, 534-544 p.Article in journal (Refereed) Published
Identifiers
urn:nbn:se:su:diva-22686 (URN)
Note
Part of urn:nbn:se:su:diva-1032Available from: 2006-05-08 Created: 2006-05-08Bibliographically approved
2. In vivo involvement of mutated initiation factor IF1 in gene expression control at the translational level
Open this publication in new window or tab >>In vivo involvement of mutated initiation factor IF1 in gene expression control at the translational level
2005 In: FEBS Letters, ISSN 0014-5793, Vol. 579, no 5, 995-1000 p.Article in journal (Refereed) Published
Identifiers
urn:nbn:se:su:diva-22687 (URN)
Note
Part of urn:nbn:se:su:diva-1032Available from: 2006-05-08 Created: 2006-05-08Bibliographically approved
3. RNA chaperone activity of translation initiation factor IF1
Open this publication in new window or tab >>RNA chaperone activity of translation initiation factor IF1
Show others...
2006 (English)In: Biochimie, ISSN 0300-9084, E-ISSN 1638-6183, Vol. 88, no 12, 1875-1882 p.Article in journal (Refereed) Published
Abstract [en]

Translation initiation factor IF1 is an indispensable protein for translation in prokaryotes. No clear function has been assigned to this factor so far. In this study we demonstrate an RNA chaperone activity of this protein both in vivo and in vitro. The chaperone assays are based on in vivo or in vitro splicing of the group I intron in the thymidylate synthase gene (td) from phage T4 and an in vitro RNA annealing assay. IF1 wild-type and mutant variants with single amino acid substitutions have been analyzed for RNA chaperone activity. Some of the IF1 mutant variants are more active as RNA chaperones than the wild-type. Furthermore, both wild-type IF1 and mutant variants bind with high affinity to RNA in a band-shift assay. It is suggested that the RNA chaperone activity of IF1 contributes to RNA rearrangements during the early phase of translation initiation.

Keyword
Escherichia coli; Initiation factor IF1; RNA chaperone; RNA folding; 30S subunit
Identifiers
urn:nbn:se:su:diva-22688 (URN)10.1016/j.biochi.2006.06.017 (DOI)
Note
Part of urn:nbn:se:su:diva-1032Available from: 2006-05-08 Created: 2006-05-08 Last updated: 2017-12-13Bibliographically approved

Open Access in DiVA

fulltext(2542 kB)2908 downloads
File information
File name FULLTEXT01.pdfFile size 2542 kBChecksum SHA-1
7d1ef89b6491879d3684987c96d0320bfd691ac76b1eb0eb595682d6dc82c4a9c25e2dcb
Type fulltextMimetype application/pdf

By organisation
Department of Genetics, Microbiology and Toxicology
Natural Sciences

Search outside of DiVA

GoogleGoogle Scholar
Total: 2908 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

isbn
urn-nbn

Altmetric score

isbn
urn-nbn
Total: 526 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf