Signal peptide hydrophobicity is finely tailored for function.
In order to titrate the dependence of individual steps in protein transport on signal peptide hydrophobicity, we have examined a series of mutants which involve replacement of the hydrophobic core segment of the Escherichia coli alkaline phosphatase signal peptide. The core regions vary in composition from 10:0 to 0:10 in the ratio of alanine to leucine residues. Thus, a nonfunctional polyalanine-containing signal peptide is titrated with the more hydrophobic residue, leucine. Analysis of this series identified a midpoint for rapid precursor processing between alanine to leucine ratios of 6:4 and 5:5 [Doud et al. (1993): Biochemistry 32:1251-1256]. Examination of precursors that are processed more slowly indicates a lower limit of signal peptide hydrophobicity that permits membrane association and translocation. Analysis of precursors that are processed rapidly defines an intermediate range of hydrophobicity that is optimum; above this level precursors become insensitive to transport inhibitors such as sodium azide and carbonyl cyanide 3-chlorophenylhydrazone (CCCP) in parallel with substantial inhibition of beta-lactamase processing. Our data indicate that there is a surprisingly narrow range of signal peptide hydrophobicity which both supports transport of the protein to which it is attached and which does not have such a high affinity for the transport pathway that it disrupts the appropriate balance of other secreted proteins.
Rusch
S L
SL
Department of Molecular and Cell Biology, University of Connecticut, Storrs 06269.
Chen
H
H
Izard
J W
JW
Kendall
D A
DA
D000409:Alanine / Q000032:analysis; D000469:Alkaline Phosphatase / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D001386:Azides / Q000494:pharmacology; D001692:Biological Transport; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004926:Escherichia coli / Q000201:enzymology; D007930:Leucine / Q000032:analysis; D011498:Protein Precursors / Q000378:metabolism; D021382:Protein Sorting Signals / Q000737:chemistry* / Q000502:physiology*; D019810:Sodium Azide; D013329:Structure-Activity Relationship; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000378:metabolism
D001386:Azides; D011498:Protein Precursors; D021382:Protein Sorting Signals; D065093:beta-Lactamase Inhibitors; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D019810:Sodium Azide; D000469:Alkaline Phosphatase; D001618:beta-Lactamases; D007930:Leucine; D000409:Alanine