Effect of Lys(-20) substitutions in alkaline phosphatase signal peptide on secretion of the enzyme

Z. N. Karamysheva, A. L. Karamyshev, V. N. Ksenzenko, M. A. Nesmeyanova

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

The effect of substitutions for positively charged Lys(-20) in the N-terminal domain of the E. coli alkaline phosphatase signal peptide on the enzyme secretion has been studied. The amber-suppressor method was used to obtain mutant alkaline phosphatases: amber mutation was introduced in the appropriate position of the alkaline phosphatase gene using oligonucleotide-directed mutagenesis, and mutant proteins were synthesized in E. coli strains producing amber-suppressor tRNAs specific for Tyr, Gly, Ala, Glu, Phe, His, Cys, and Pro. All the mutant proteins can be translocated across the cytoplasmic membrane and assembled in the periplasm to active enzyme molecules. However, certain amino acid substitutions reduce the rate of protein maturation. Their effect depends not only on the charge of the amino acid residue, but also on its nature. Thus, introduction of positively charged His and the polar uncharged Tyr has no significant effect, while introduction of negatively charged Glu and the hydrophobic Ala, Phe, and Pro, as well as Gly and Cys, has an inhibiting action. The data show the important role of the primary structure of the signal peptide N-terminus in secretion.

Original languageEnglish (US)
Pages (from-to)541-548
Number of pages8
JournalBiochemistry (Moscow)
Volume61
Issue number4
StatePublished - 1996

Fingerprint

Protein Sorting Signals
Amber
Alkaline Phosphatase
Substitution reactions
Mutant Proteins
Escherichia coli
Enzymes
RNA, Transfer, Tyr
Amino Acids
Periplasm
Mutagenesis
Amino Acid Substitution
Site-Directed Mutagenesis
Transfer RNA
Oligonucleotides
Genes
Cell Membrane
Membranes
Mutation
Molecules

Keywords

  • Alkaline phosphatase
  • Amino acid substitutions
  • E. coli
  • Protein secretion
  • Signal peptide
  • Suppressor tRNA

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

Cite this

Karamysheva, Z. N., Karamyshev, A. L., Ksenzenko, V. N., & Nesmeyanova, M. A. (1996). Effect of Lys(-20) substitutions in alkaline phosphatase signal peptide on secretion of the enzyme. Biochemistry (Moscow), 61(4), 541-548.

Effect of Lys(-20) substitutions in alkaline phosphatase signal peptide on secretion of the enzyme. / Karamysheva, Z. N.; Karamyshev, A. L.; Ksenzenko, V. N.; Nesmeyanova, M. A.

In: Biochemistry (Moscow), Vol. 61, No. 4, 1996, p. 541-548.

Research output: Contribution to journalArticle

Karamysheva, ZN, Karamyshev, AL, Ksenzenko, VN & Nesmeyanova, MA 1996, 'Effect of Lys(-20) substitutions in alkaline phosphatase signal peptide on secretion of the enzyme', Biochemistry (Moscow), vol. 61, no. 4, pp. 541-548.
Karamysheva ZN, Karamyshev AL, Ksenzenko VN, Nesmeyanova MA. Effect of Lys(-20) substitutions in alkaline phosphatase signal peptide on secretion of the enzyme. Biochemistry (Moscow). 1996;61(4):541-548.
Karamysheva, Z. N. ; Karamyshev, A. L. ; Ksenzenko, V. N. ; Nesmeyanova, M. A. / Effect of Lys(-20) substitutions in alkaline phosphatase signal peptide on secretion of the enzyme. In: Biochemistry (Moscow). 1996 ; Vol. 61, No. 4. pp. 541-548.
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