A comparative analysis of single- and multiple-residue substitutions in the alkaline phosphatase signal peptide.
The alkaline phosphatase signal peptide participates in transport of the enzyme to the periplasmic space of Escherichia coli. The signal sequence, like that of other signal peptides, is composed of a polar amino-terminal segment, a central region rich in hydrophobic residues and a carboxy-terminal region recognized by signal peptidase. We have previously shown that an alkaline phosphatase signal peptide mutant containing a polyleucine core region functions efficiently in transport of the enzyme [D. A. Kendall, S. C. Bock, and E. T. Kaiser (1986) Nature 321, 706-708]. In this study, some of the amino acid changes involved in the polyleucine sequence are examined individually. A Phe to Leu substitution as the sole change results in impaired transport properties in contrast to when it is combined with three other amino acid changes in the polyleucine-containing sequence. A mutant with a Pro to Leu substitution in the hydrophobic core region is comparable to wild type while the same type of substitution (Pro to Leu) in the carboxy-terminal segment results in substantial accumulation of the mutant precursor. Finally, introduction of a basic residue into the hydrophobic segment (Leu to Arg substitution) results in a complete export block. These results exemplify the spectrum of properties produced by individual residue changes and suggest there is some interplay between hydrophobicity and conformation for signal peptide function.
Laboratory of Bioorganic Chemistry and Biochemistry, Rockefeller University, New York, New York 10021.
D A
DA
Kendall
S K
SK
Doud
E T
ET
Kaiser
D000469:Alkaline Phosphatase / Q000378:metabolism*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D008969:Molecular Sequence Data; D009154:Mutation; D021382:Protein Sorting Signals / Q000235:genetics / Q000378:metabolism*
D000596:Amino Acids; D021382:Protein Sorting Signals; D000469:Alkaline Phosphatase