Protein translocation across the endoplasmic reticulum membrane: identification by photocross-linking of a 39-kD integral membrane glycoprotein as part of a putative translocation tunnel.
The molecular environment of secretory proteins during translocation across the ER membrane was examined by photocross-linking. Nascent preprolactin chains of various lengths, synthesized by in vitro translation of truncated messenger RNAs in the presence of N epsilon-(5-azido-2-nitrobenzoyl)-Lys-tRNA, signal recognition particle, and microsomal membranes, were used to position photoreactive probes at various locations within the membrane. Upon photolysis, each nascent chain species was cross-linked to an integral membrane glycoprotein with a deduced mass of 39 kD (mp39) via photoreactive lysines located in either the signal sequence or the mature prolactin sequence. Thus, different portions of the nascent preprolactin chain are in close proximity to the same membrane protein during the course of translocation, and mp39 therefore appears to be part of the translocon, the specific site of protein translocation across the ER membrane. The similarity of the molecular and cross-linking properties of mp39 and the glyco-protein previously identified as a signal sequence receptor (Wiedmann, M., T. V. Kurzchalia, E. Hartmann, and T. A. Rapoport. 1987. Nature [Lond.]. 328: 830-833) suggests that these two proteins may be identical. Our data indicate, however, that mp39 does not (or not only) function as a signal sequence receptor, but rather may be part of a putative translocation tunnel.
Krieg
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UC
Department of Chemistry and Biochemistry, University of Oklahoma, Norman 73019.
Johnson
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AE
Walter
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P
D003432:Cross-Linking Reagents / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008861:Microsomes / Q000378:metabolism; D008970:Molecular Weight; D010782:Photolysis; D011388:Prolactin / Q000235:genetics; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000235:genetics; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics
D003432:Cross-Linking Reagents; D008562:Membrane Glycoproteins; D011498:Protein Precursors; D012333:RNA, Messenger; C017671:preprolactin; D011388:Prolactin