In vitro selection of GTP-binding proteins by block shuffling of estrogen-receptor fragments

Toru Tsuji, Michiko Onimaru, Nobuhide Doi, Etsuko Miyamoto-Sato, Hideaki Takashima, Hiroshi Yanagawa

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

To what extent has alternative splicing contributed to the evolution of protein-function diversity? We previously constructed a pool of block-deletion mutants of the human estrogen receptor α ligand binding domain by random multi-recombinant PCR. Here we performed iterative in vitro selection of GTP-binding proteins by using the library of mRNA-displayed proteins and GTP-affinity chromatography combined with quantitative real-time PCR. We obtained a novel GTP-binding protein with moderate affinity and substrate-specificity. The results of our in vitro simulation imply that alternative splicing may have contributed substantially to the diversification of protein function during evolution.

Original languageEnglish
Pages (from-to)689-693
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume390
Issue number3
DOIs
Publication statusPublished - 2009 Dec 18

Fingerprint

GTP-Binding Proteins
Estrogen Receptors
Alternative Splicing
Affinity chromatography
Proteins
Substrate Specificity
Guanosine Triphosphate
Affinity Chromatography
Real-Time Polymerase Chain Reaction
Ligands
Polymerase Chain Reaction
Messenger RNA
Substrates
In Vitro Techniques

Keywords

  • Alternative splicing
  • Combinatorial protein library
  • In vitro selection
  • mRNA display
  • Protein evolution
  • Synthetic biology

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Cell Biology
  • Molecular Biology

Cite this

In vitro selection of GTP-binding proteins by block shuffling of estrogen-receptor fragments. / Tsuji, Toru; Onimaru, Michiko; Doi, Nobuhide; Miyamoto-Sato, Etsuko; Takashima, Hideaki; Yanagawa, Hiroshi.

In: Biochemical and Biophysical Research Communications, Vol. 390, No. 3, 18.12.2009, p. 689-693.

Research output: Contribution to journalArticle

Tsuji, Toru ; Onimaru, Michiko ; Doi, Nobuhide ; Miyamoto-Sato, Etsuko ; Takashima, Hideaki ; Yanagawa, Hiroshi. / In vitro selection of GTP-binding proteins by block shuffling of estrogen-receptor fragments. In: Biochemical and Biophysical Research Communications. 2009 ; Vol. 390, No. 3. pp. 689-693.
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