N-terminal basic amino acids are not required for translocation and processing of preproparathyroid hormone.
An N-terminal deletion mutant of preproparathyroid hormone that contains a single basic amino acid, lysine, in the N-terminal domain of the signal peptide is translocated across the endoplasmic reticulum membrane similarly to intact preproparathyroid hormone. To examine the function of charged residues preceeding the hydrophobic core, the lysine was replaced by an uncharged (methionine) or negatively charged (glutamic acid) amino acid. The translocational activity of the mutant signal peptides was assayed in a reticulocyte lysate system containing chicken oviduct microsomal membranes. Altering the net charge of the N-terminal domain did not abolish signal sequence activity, although the efficiency of translocation was decreased for the mutant with a glutamic acid substitution. Posttranslational, ribosome independent, translocation was observed for all the mutants tested, with the same dependence on N-terminal charge but with much lower efficiency than cotranslational translocation. These studies show that the presence of basic amino acids in the N-terminal domain of a eukaryotic signal sequence is not required for its activity.
Szczesna-Skorupa
E
E
Department of Physiology and Biophysics, University of Illinois, Urbana-Champaign 61801.
Kemper
B
B
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002645:Chickens; D004563:Electrochemistry; D005260:Female; D005971:Glutamates; D018698:Glutamic Acid; D007425:Intracellular Membranes / Q000378:metabolism; D008715:Methionine; D008861:Microsomes / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010057:Oviducts / Q000378:metabolism; D010281:Parathyroid Hormone / Q000235:genetics / Q000378:metabolism*; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000502:physiology*; D012156:Reticulocytes / Q000378:metabolism; D013329:Structure-Activity Relationship
D005971:Glutamates; D010281:Parathyroid Hormone; D011498:Protein Precursors; D021382:Protein Sorting Signals; D018698:Glutamic Acid; C019056:preproparathormone; D008715:Methionine