Expression of biologically active rat apolipoprotein AIV in Escherichia coli

Min Liu, Nick Maiorano, Ling Shen, Kevin Pearson, Daisuke Tajima, Dian Ming Zhang, Stephen C. Woods, Randy J. Seeley, W. Sean Davidson, Patrick Tso

Research output: Contribution to journalArticlepeer-review

26 Scopus citations

Abstract

Rat apolipoprotein AIV (apo AIV) is a 43-kDa intestinal apolipoprotein that is important in lipid metabolism and the suppression of food intake. In this study, a full-length rat apo AIV was expressed in Escherichia coli and purified in a bioactive form. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometric analysis revealed that the isolated recombinant protein has a molecular mass of approximately 43 kDa, similar to that of natural rat apo AIV. Immunoblot analysis and N-terminal amino acid sequencing confirmed the identity of the recombinant apo AIV protein as natural rat apo AIV. The recombinant protein was functional in lipoprotein binding assays. Biological activity was assessed behaviorally in that the recombinant protein suppressed food intake of fasted rats comparably to natural rat apo AIV. Neither native nor recombinant apo AIV elicited a conditioned taste aversion (CTA) at doses that suppress feeding. These results indicate that the recombinant apo AIV is structurally and functionally indistinguishable from rat natural apo AIV, making this overexpression and purification scheme a powerful tool for future structure and function studies.

Original languageEnglish
Pages (from-to)149-155
Number of pages7
JournalPhysiology and Behavior
Volume78
Issue number1
DOIs
StatePublished - Jan 2003

Bibliographical note

Funding Information:
The authors gratefully acknowledge the help of Dr. David Hui in providing the rat apo AIV cDNA. We also wish to thank Dr. Feng Wang for performing electrospray mass spectrometry and N-terminal sequence of the recombinant protein. This work was supported by research grants from National Institutes of Health DK54890, DK17844, DK56863, HL62542-01, DK54504, DK56910 and DK53444, as well as supported by Proctor and Gamble.

Keywords

  • Apolipoprotein A-IV
  • Escherichia coli
  • Food intake
  • Recombinant expression

ASJC Scopus subject areas

  • Experimental and Cognitive Psychology
  • Behavioral Neuroscience

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