Studies of the type I cellular retinoic acid-binding protein mutants and their biological activities

Li-Na Wei, Liming Chang, Xinli Hu

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

We have mutated the type I cellular retinoic acid binding protein (CRABP-I), individually at the Arg131 (into Ala) and the Tyr133 (into Phe) residues which have been predicted to make direct contact with retinoic acid (RA) based upon previous structural studies. The RA-binding affinities of these mutants are examined and their biological effects on RA induction of reporter genes are determined. The R131A mutation drastically affects its ligand-binding property, but the Y133F mutation has little effect. By using an RA-inducible reporter, it is found that the wild type CRABP-I exerts biphasic effects on RA induction of the reporter. The early (at 12 h) effect is to enhance RA induction, whereas the delayed (at 24 h) effect is to suppress RA induction. In consistence with their RA binding property, the R131A mutant loses both its early and delayed biological activities, whereas the Y133F mutant remains as effective as the wild type. It is concluded that CRABP-I over-expression exerts biphasic effects on RA-mediated gene expression, and that Arg131, but not Tyr133, is essential for a high RA-binding affinity of this protein as well as its biological activity.

Original languageEnglish (US)
Pages (from-to)69-76
Number of pages8
JournalMolecular and cellular biochemistry
Volume200
Issue number1-2
DOIs
StatePublished - 1999

Bibliographical note

Funding Information:
We thank Drs. L. Banaszak and J. Thompson for providing the structural information about CRABP-I, discussion about this study and their generosity in sharing equipment. We thank Dr. H-C. Lee for sharing the fluorometer in our fluorescent titration experiments. We also thank core B of the program project (DA08131) for oligonucleotide synthesis. This work was supported by DK46866 of NIH, 98-35200-6264 of USDA and, in part, by the U. Minn. Graduate School Sponsored Initiative in Structural Biology to LNW.

Keywords

  • CRABP-I
  • Mutagenesis
  • RA binding
  • RA induction

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