title: Identification of TMEM45B as a protein clearly showing thermal aggregation in SDS-PAGE gels and dissection of its amino acid sequence responsible for this aggregation.
abstract: SDS-PAGE is one of the most powerful experimental techniques used for the separation of proteins, and most proteins are separated according to their molecular size by this technique. However, exceptional proteins showing abnormal behavior in SDS-PAGE gels are known to exist. Thermal aggregation, rarely observed with membrane proteins, is one of the exceptional behaviors of proteins during SDS-PAGE, but detailed characterization of this aggregation has not yet been achieved. In the present study, we found that a putative membrane protein, TMEM45B, very clearly showed properties of thermal aggregation when it was expressed in COS7 cells and subjected to SDS-PAGE. We dissected the region of TMEM45B responsible for this aggregation, and found that of the seven putative transmembrane domains, a region comprising the 4th to 7th ones was essential for the thermal aggregation properties. We also demonstrated that these transmembrane domains, 4th to 7th, of TMEM45B conferred thermal aggregation properties on other proteins, by fusing this amino acid sequence to target proteins. The molecular mechanism causing thermal aggregation by TMEM45B is still uncertain, but TMEM45B could be utilized as a nice model to show clear thermal aggregation in SDS-PAGE gels.
authors.affiliation: Faculty of Pharmaceutical Sciences, University of Tokushima, Shomachi-1, Tokushima 770-8505, Japan.
authors.forename: Naoto
authors.initials: N
authors.lastname: Okada
authors.forename: Takenori
authors.initials: T
authors.lastname: Yamamoto
authors.forename: Masahiro
authors.initials: M
authors.lastname: Watanabe
authors.forename: Yuuya
authors.initials: Y
authors.lastname: Yoshimura
authors.forename: Eriko
authors.initials: E
authors.lastname: Obana
authors.forename: Naoshi
authors.initials: N
authors.lastname: Yamazaki
authors.forename: Kazuyoshi
authors.initials: K
authors.lastname: Kawazoe
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Shinohara
authors.forename: Kazuo
authors.initials: K
authors.lastname: Minakuchi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002522:Chlorocebus aethiops; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006358:Hot Temperature; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D011489:Protein Denaturation; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D011994:Recombinant Proteins