Mass-production of human ACAT-1 and ACAT-2 to screen isoform-specific inhibitor: A different substrate specificity and inhibitory regulation

Kyung Hyun Cho, Sojin An, Woo Song Lee, Young Ki Paik, Young Kook Kim, Tae Sook Jeong

Research output: Contribution to journalArticlepeer-review

42 Citations (Scopus)


Recently, acyl-CoA:cholesterol acyltransferase was found to be present as two isoforms, ACAT-1 and ACAT-2, in mammalian tissues with different metabolic functions and tissue-specific locations. In this study, the isoforms were mass-produced individually from insect cells to establish a more sensitive and reliable screening method for specific inhibitors against each isoform. The expressed hACAT-1 and hACAT-2 appeared as a 50kDa- and a 46kDa-band on SDS-PAGE, respectively, from Hi5 cells and they preferred to exist in oligomeric form, from dimer to tetramer, during the purification process. They also exhibited an approximate 3.4 to 3.7-fold increase in activities when compared to rat liver microsomal fractions at the same protein concentration. Known ACAT inhibitors, pyripyropene A, oleic acid anilide, and diethyl pyrocarbonate, were tested to evaluate the inhibitory specificity and sensitivity of the expressed enzymes. Interestingly, pyripyropene A inhibited only the hACAT-2 fraction with IC 50=0.64μM but not the hACAT-1 fraction; whereas the fatty acid anilide did not show a significant difference in inhibitory activity with either hACAT-1 or hACAT-2. Furthermore, cholesterol was more rapidly utilized by hACAT-1, but hACAT-2 esterified other cholic acid derivatives more efficiently. These results suggest that the specificity of each substrate and inhibitor was highly different, depending on each isoform from the viewpoint of the regulatory site and the substrate binding site location.

Original languageEnglish
Pages (from-to)864-872
Number of pages9
JournalBiochemical and Biophysical Research Communications
Issue number4
Publication statusPublished - 2003 Oct 3

Bibliographical note

Funding Information:
This work is supported by a grant from the Korea Health 21 R&D project, Ministry of Health & Welfare, Republic of Korea (No. 02-PJ1-PG10-20999-0001).

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology


Dive into the research topics of 'Mass-production of human ACAT-1 and ACAT-2 to screen isoform-specific inhibitor: A different substrate specificity and inhibitory regulation'. Together they form a unique fingerprint.

Cite this