122nd General Meeting of the KCS

Type Oral Presentation
Area Oral Presentation of Young Analytical Chemists I
Room No. Room 321
Time THU 10:30-:
Code ANAL1.O-16
Subject Optimal purification method for expression of human melanocortin-4 receptor
Authors Minseon Kim, Soyeon Jo1, Ji Sun Kim 2, Yongae Kim2,*
Department of chemistry, Hankuk University of Foreign Studies, Korea
1chemistry, Hankuk University of Foreign Studies, Korea
2Department of Chemistry, Hankuk University of Foreign Studies, Korea
Abstract The human transmembrane protein (hTMP) is the type of integral membrane proteins and exists as a signal transduction, intercellular communication, and ion channels, etc. Thus, it has various functions in the biological membrane. In order to demonstrate the function of the transmembrane proteins, the protein should be purified to identify the structure because function is related to structure closely. However, since the transmembrane protein is most composed of hydrophobic amino acids and is surrounded by the lipid, expression and purification of transmembrane protein are not easy. The transmembrane protein, which plays a biologically important role, is associated with the onset of many diseases. If a mutation occurs in human melanocortin-4 receptor (hMC4R), one of the transmembrane proteins, cause eating disorder and obesity. Asparagine-substituted mutants in aspartic acid, the 90th amino acid in the second transmembrane protein (TM2), were found in the patients with early onset obesity. It was thought that the loss of function was caused by structural changes of hMC4R due to the D90N mutation. Therefore, we examined the structural differences between wild-type hMC4R-TM2 (wt-hMC4R-TM2) and mutant hMC4R-TM2 (m-hMC4R-TM2) after expression and purification of two proteins. In the experimental process, SDS was used as a detergent when separating proteins using FPLC to separate them from impurities using hydrophobicity of target proteins. Afterwards, purified high yield target proteins were obtained by using SDS removal method, and the final structure was confirmed by various spectroscopic methods like MS, CD, solution NMR spectroscopy, and solid-state NMR spectroscopy.
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