title: Proline in transmembrane domain of type II protein DPP-IV governs its translocation behavior through endoplasmic reticulum.
abstract: A transmembrane domain (TMD) at the N-terminus of a membrane protein is a signal sequence that targets the protein to the endoplasmic reticulum (ER) membrane. Proline is found more frequently in TM helices compared to water-soluble helices. To investigate the effects of proline on protein translocation and integration in mammalian cells, we made proline substitutions throughout the TMD of dipeptidyl peptidase IV, a type II membrane protease with a single TMD at its N-terminus. The proteins were expressed and their capacities for targeting and integrating into the membrane were measured in both mammalian cells and in vitro translation systems. Three proline substitutions in the central region of the TMD resulted in various defects in membrane targeting and/or integration. The replacement of proline with other amino acids of similar hydrophobicity rescued both the translocation and anchoring defects of all three proline mutants, indicating that conformational change caused by proline is a determining factor. Increasing hydrophobicity of the TMD by replacing other residues with more hydrophobic residues also effectively reversed the translocation and integration defects. Intriguingly, increasing hydrophobicity at the C-terminal end of the TMD rescued much more effectively than it did at the N-terminal end. Thus, the effect of proline on translocation and integration of the TMD is not determined solely by its conformation and hydrophobicity, but also by the location of proline in the TMD, the location of highly hydrophobic residues, and the relative position of the proline to other proline residues in the TMD.
authors.affiliation: Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Miaoli County 350, Taiwan, ROC.
authors.forename: Kuei-Min
authors.initials: KM
authors.lastname: Chung
authors.forename: Chih-Hsiang
authors.initials: CH
authors.lastname: Huang
authors.forename: Jai-Hong
authors.initials: JH
authors.lastname: Cheng
authors.forename: Cheng-Han
authors.initials: CH
authors.lastname: Tsai
authors.forename: Ching-Shu
authors.initials: CS
authors.lastname: Suen
authors.forename: Ming-Jing
authors.initials: MJ
authors.lastname: Hwang
authors.forename: Xin
authors.initials: X
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D018819:Dipeptidyl Peptidase 4 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004285:Dogs; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D011392:Proline / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology
chemical_list: D011392:Proline; C042807:DPP4 protein, human; D018819:Dipeptidyl Peptidase 4