We have previously described a novel mutant of human preproapolipoprotein A-II (pre(Δpro)apoA-II) in which the wild-type 18-amino acid-long signal sequence (Gly(18↓)) was functionally redefined to 20 amino acids in length (Ala(20↓)). We have used this mutant as a model preprotein to probe the substrate specificity of eukaryotic signal peptidase. Site-saturation mutagenesis was performed resulting in the substitution of 13 different amino acids (acidic, basic, aromatic, hydrophobic, and small-neutral) for Ala20 (or position -1). The effects of these substitutions were assessed using an in vitro transcription/translation/microsomal membrane processing system. NH2-terminal sequence analysis of the 13 mutant proteins demonstrated that amino acids which occupy position -1 in a signal peptide are critical in establishing a good context for signal peptidase cleavage and that two or more potential sites of cleavage may compete for recognition by signal peptidase.
|Original language||English (US)|
|Number of pages||9|
|Journal||Journal of Biological Chemistry|
|State||Published - 1988|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology