title: The long D-stem of the selenocysteine tRNA provides resilience at the expense of maximal function.
abstract: BACKGROUND
  The selenocysteine tRNA (tRNASec) has a uniquely long D-stem containing 6 base pairs.
  
  
  RESULTS
  The extended D-stem is not essential for function but is required for stability.
  
  
  CONCLUSIONS
  Enhanced secondary structure in selenocysteine tRNA compensates for the absence of canonical tertiary interactions.
  
  
  CONCLUSIONS
  The flexibility due to the absence of tertiary interactions is required for tRNASec function, whereas the enhanced secondary structure compensates for the decreased stability. The D-stem of the selenocysteine tRNA (tRNA(Sec)) contains 2 additional base pairs, which replace tertiary interactions 8-14 and 15-48 universally present in all other cytosolic tRNAs. To study the role of these additional base pairs in the tRNA(Sec) function, we used the instant evolution approach. In vivo screening of six combinatorial gene libraries provided 158 functional variants of the Escherichia coli tRNA(Sec). Analysis of these variants showed that the additional base pairs in the D-stem were not required for the tRNA(Sec) function. Moreover, at lower temperatures, these base pairs notably harmed the tRNA(Sec) activity. However, at elevated temperatures, these base pairs became essential as they made the tRNA structure more stable. The alternative way to stabilize the structure through formation of the standard tertiary interactions was not an option for tRNA(Sec) variants, which suggests that the absence of these interactions and the resulting flexibility of the tertiary structure are essential for tRNA(Sec) function.
authors.affiliation: Department of Biochemistry, Université de Montréal, Succursale Centre-ville, Montréal, Québec H3C 3J7, Canada.
authors.forename: Tetsu M
authors.initials: TM
authors.lastname: Ishii
authors.forename: Natalia
authors.initials: N
authors.lastname: Kotlova
authors.forename: Franck
authors.initials: F
authors.lastname: Tapsoba
authors.forename: Sergey V
authors.initials: SV
authors.lastname: Steinberg
mesh_terms: D001483:Base Sequence; D019020:Directed Molecular Evolution; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D055029:Inverted Repeat Sequences; D008969:Molecular Sequence Data; D020871:RNA Stability; D012329:RNA, Bacterial / Q000235:genetics* / Q000502:physiology; D012345:RNA, Transfer, Amino Acid-Specific / Q000235:genetics* / Q000502:physiology; D017279:Selenocysteine; D046249:Transfer RNA Aminoacylation
chemical_list: D012329:RNA, Bacterial; D012345:RNA, Transfer, Amino Acid-Specific; C036928:tRNA, selenocysteine-; D017279:Selenocysteine
keywords: Combinatorial Library; In Vivo Screening; Instant Evolution; RNA Folding; RNA Structure; Ribosome Function; Selenocysteine; Tertiary Structure; Transfer RNA (tRNA)