Physicochemical properties analysis and prokaryotic expression of human cholesterol ester transfer protein
LI Shuxuan
LI Jing
FENG Zhangyi
BI Yancheng
ZHANG Ming
QIU Hongquan
LI Chunyan
ZHANG Defu
Abstract：Cholesteryl ester transfer protein(CETP)plays a critical regulatory role in the pathogenesis of hyperlipidemia.It facilitates the transfer of cholesteryl esters from high-density lipoprotein(HDL)to very-low-density lipoprotein(VLDL)and low-density lipoprotein(LDL),thereby reducing cardioprotective HDL cholesterol levels and elevating pro-atherogenic LDL cholesterol levels,which ultimately promotes the development of hyperlipidemia.To enable prokaryotic expression,the coding sequence(CDS)of CETP was retrieved from the GenBank database.Following bioinformatic analysis and codon optimization,a pCold-CETP prokaryotic expression vector was constructed,and the target protein was purified via nickel-affinity chromatography.Bioinformatic analysis revealed that the CETP gene CDS encodes a 493-amino acid protein with a predicted molecular weight of approximately 54 ku.The protein was predicted to be stable(instability index:32.68)and hydrophilic.After codon optimization,the average guanine and cytosine(GC)content of the CETP gene was reduced from 54.7%to 51.3%,and unfavorable secondary structures were eliminated,significantly enhancing its expression efficiency in Escherichia coli.Under induction conditions of 37℃and 10 mmol/L isopropyl β-D-1-thiogalactopyranoside(IPTG)overnight,the CETP protein was predominantly expressed in inclusion bodies.The purified protein showed a distinct band on SDS-PAGE,with a final concentration of 13.48 mg/mL.In conclusion,this study successfully achieved high-yield expression of the CETP protein in a prokaryotic system.The significant improvement in protein production,accomplished through codon optimization and induction condition refinement,provides a solid foundation for future functional studies of CETP and the development of its inhibitors.
Keywords：cholesterol ester transfer proteinbioinformaticsprokaryotic expressionrecombinant proteinprotein purification
Publication Date：2025-12-15
Online Publishing Date：2026-04-01(First online date of this platform, not the publication date of the document)
Pages：13( 251-263 )