Cloning and characterization of a novel apolipoprotein gene, apolipoprotein AV, in tree shrews

Mol Biol Rep. 2013 Sep;40(9):5429-38. doi: 10.1007/s11033-013-2641-0. Epub 2013 May 17.

Abstract

Apolipoprotein AV (apoAV) modulates plasma triglyceride levels, which is an independent risk factor for cardiovascular disease. ApoAV is also involved in atherosclerosis lesion formation. In order to systematically evaluate the apolipoprotein-related gene profile in tree shrew, a model for its insusceptibility to atherosclerosis, we performed apoAV cloning and characterization. The full-length cDNA of apoAV was identified using SMART-RACE. ApoAV cDNA sequence revealed two transcripts, 1,948 and 1,397 base pairs, due to alternative polyadenylation. These two transcripts share the same open reading frame (ORF), which encodes a 369-amino acid protein with high identity to human apoAV (75 %), including a 23-amino acid N-terminal signal peptide. ApoAV is expressed exclusively in the liver. Mature apoAV was expressed in E. coli BL21(DE3) and purified by Ni-chelated resin. Lipoprotein lipase activity was significantly stimulated by this recombinant protein. The full-length ORF of apoAV was cloned into pDsRed-monomer-N1 vector with a red fluorescent protein tag and was primarily localized in cytoplasm of hepG2 cells. The successful cloning, expression and localization of apoAV in tree shrew has laid down the foundation for further investigation on its structure and functions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Apolipoproteins / genetics*
  • Apolipoproteins / metabolism
  • Base Sequence
  • Cloning, Molecular
  • Computational Biology
  • DNA Primers / genetics
  • DNA, Complementary / genetics
  • Escherichia coli
  • Gene Expression Profiling
  • Genetic Vectors / genetics
  • Immunoblotting
  • Liver / metabolism*
  • Luminescent Proteins
  • Molecular Sequence Data
  • Open Reading Frames / genetics
  • Recombinant Proteins / genetics*
  • Recombinant Proteins / metabolism
  • Sequence Analysis, DNA
  • Tupaiidae / genetics*
  • Tupaiidae / metabolism

Substances

  • Apolipoproteins
  • DNA Primers
  • DNA, Complementary
  • Luminescent Proteins
  • Recombinant Proteins
  • red fluorescent protein