Selenocysteine tRNA [Ser]Sec, the Central Component of Selenoprotein Biosynthesis: Isolation, Identification, Modification, and Sequencing

Methods Mol Biol. 2018;1661:43-60. doi: 10.1007/978-1-4939-7258-6_4.

Abstract

The selenocysteine (Sec) tRNA[Ser]Sec population consists of two isoforms that differ from each other by a single 2'-O-methylribosyl moiety at position 34 (Um34). These two isoforms, which are encoded in a single gene, Trsp, and modified posttranscriptionally, are involved individually in the synthesis of two subclasses of selenoproteins, designated housekeeping and stress-related selenoproteins. Techniques used in obtaining these isoforms for their characterization include extraction of RNA from mammalian cells and tissues, purifying the tRNA[Ser]Sec population by one or more procedures, and finally resolving the two isoforms from each other. Since some of the older techniques for isolating tRNA[Ser]Sec and resolving the isoforms are used in only a few laboratories, these procedures will be discussed briefly and references provided for more detailed information, while the more recently developed procedures are discussed in detail. In addition, a novel technique that was developed in sequencing tRNA[Ser]Sec for identifying their occurrence in other organisms is also presented.

Keywords: Chromatography; Gel electrophoresis; Selenium; Selenocysteine tRNA; Selenocysteine tRNA detection; Selenocysteine tRNA gene modification.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Intramural

MeSH terms

  • Animals
  • Blotting, Northern
  • Chromatography, Affinity
  • Chromatography, Reverse-Phase
  • Humans
  • Isotope Labeling
  • Nucleic Acid Conformation
  • Protein Biosynthesis
  • RNA, Transfer, Amino Acid-Specific / chemistry
  • RNA, Transfer, Amino Acid-Specific / genetics*
  • Selenium Radioisotopes
  • Selenoproteins / chemistry
  • Selenoproteins / genetics*
  • Selenoproteins / isolation & purification
  • Sequence Analysis, RNA

Substances

  • RNA, Transfer, Amino Acid-Specific
  • Selenium Radioisotopes
  • Selenoproteins
  • tRNA, selenocysteine-