Stop-transfer activity of hydrophobic sequences depends on the translation system.
Signal and stop-transfer sequences are the known determinants involved in topogenesis of integral membrane proteins. To study the characteristics of stop-transfer sequences, artificial proteins have been created on the DNA level based on the cDNA of the asialoglycoprotein receptor H1. Its internal signal/anchor domain initiates translocation of the downstream sequence across the endoplasmic reticulum membrane. The ability of several hydrophobic sequences inserted into the translocating polypeptide to stop further transfer was analyzed by translation of the fusion proteins using the wheat germ extract and rabbit reticulocyte lysate systems with dog pancreas microsomes. We discovered that some of the sequences behave differently with respect to translocation across the membrane depending on the translation system. Expression of one of the fusion proteins in fibroblasts showed that the reticulocyte lysate system reflects more closely the in vivo situation than the wheat germ system. Our results suggest that in a homologous system the translating ribosomes interact with the translocation machinery and influence the termination of polypeptide transfer by hydrophobic sequences.
Spiess
M
M
Department of Biochemistry, University of Basel, Switzerland.
Handschin
C
C
Baker
K P
KP
D000595:Amino Acid Sequence; D037263:Asialoglycoprotein Receptor; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D014176:Protein Biosynthesis; D011487:Protein Conformation; D021382:Protein Sorting Signals / Q000235:genetics*; D011971:Receptors, Immunologic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D014158:Transcription, Genetic
D037263:Asialoglycoprotein Receptor; D021382:Protein Sorting Signals; D011971:Receptors, Immunologic; D011993:Recombinant Fusion Proteins; D004247:DNA