Home LiteratureArticle Details
PMID: 2471921 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Modification of length, hydrophobic properties and electric charge of Bacillus subtilis alpha-amylase signal peptide and their different effects on the production of secretory proteins in B. subtilis and Escherichia coli cells.

Molecular & general genetics : MGG ·Vol. 216 ·No. 1 ·1989-03-00 ·Pages 1-9

Nakamura K, Fujita Y, Itoh Y, Yamane K

Abstract

Bacillus subtilis alpha-amylase signal peptide, which consists of 33 amino acids, is functional in Escherichia coli cells. Lysine, glutamic acid, leucine, leucyl-leucine, or leucyl-leucyl-leucine was inserted between positions 28 and 29 of the alpha-amylase signal peptide using site directed mutagenesis. DNAs encoding the wild-type and modified signal peptides were then fused in-frame to DNAs encoding the mature regions of the beta-lactamase of pBR322 and a thermostable alpha-amylase. The secretion of beta-lactamase in E. coli cells was more inhibited by the modified signal peptides than that in B. subtilis cells, although the degree of inhibition varied and the inhibitory effect of each signal peptide was found to be similar in the two strains. In contrast, the difference in the inhibitory effect of each modified signal peptide was no longer detected in the case of the production of thermostable alpha-amylase, except for the insertion of glutamic acid. Nearly 50% of thermostable alpha-amylase in the precursor form was accumulated in the intracellular fraction of E. coli cells containing the DNAs for the modified signal peptides. The insertion of glutamic acid inhibited the secretion of the two enzymes in both B. subtilis and E. coli cells.

MeSH Terms
Amino Acid Sequence Bacillus subtilis/genetics,metabolism Bacterial Proteins/biosynthesis Base Sequence DNA, Bacterial/genetics Escherichia coli/metabolism Molecular Sequence Data Plasmids Protein Sorting Signals/genetics,metabolism RNA, Bacterial/genetics RNA, Messenger/genetics alpha-Amylases/genetics,metabolism beta-Lactamases/biosynthesis
Chemicals
Bacterial Proteins DNA, Bacterial Protein Sorting Signals RNA, Bacterial RNA, Messenger alpha-Amylases beta-Lactamases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Nakamura K
Institute of Biological Sciences, University of Tsukuba, Ibaraki, Japan.
Fujita Y
Itoh Y
Yamane K
References (29)
29 references, click to expand
  1. Protein processing to form extracellular thermostable alpha-amylases from a gene fused in a Bacillus subtilis secretion vector.
    J Gen Microbiol. 1987 Nov;133(11):3271-7 PMID: 3128640
  2. Cleavage of structural proteins during the assembly of the head of bacteriophage T4.
    Nature. 1970 Aug 15;227(5259):680-5 PMID: 5432063
  3. Alpha-amylase genes (amyR2 and amyE+) from an alpha-amylase-hyperproducing Bacillus subtilis strain: molecular cloning and nucleotide sequences.
    J Bacteriol. 1983 Oct;156(1):327-37 PMID: 6413492
  4. NH2-terminal processing of Bacillus subtilis alpha-amylase.
    J Biol Chem. 1988 Aug 15;263(23):11548-53 PMID: 3136160
  5. Sequence analysis of mutations that prevent export of lambda receptor, an Escherichia coli outer membrane protein.
    Nature. 1980 May 8;285(5760):82-5 PMID: 6445509
  6. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose.
    Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201-5 PMID: 6159641
  7. Compilation of published signal sequences.
    Nucleic Acids Res. 1984 Jul 11;12(13):5145-64 PMID: 6379599
  8. The differential effect on two hybrid proteins of deletion mutations within the hydrophobic region of the Escherichia coli OmpA signal peptide.
    J Biol Chem. 1987 Feb 5;262(4):1716-9 PMID: 3543010
  9. Mutations that alter the signal sequence of alkaline phosphatase in Escherichia coli.
    J Bacteriol. 1983 Apr;154(1):366-74 PMID: 6339478
  10. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments.
    Gene. 1982 Oct;19(3):269-76 PMID: 6295880
  11. Transformation of Salmonella typhimurium by plasmid deoxyribonucleic acid.
    J Bacteriol. 1974 Sep;119(3):1072-4 PMID: 4605400
  12. Secretion activities of Bacillus subtilis alpha-amylase signal peptides of different lengths in Escherichia coli cells.
    Biochem Biophys Res Commun. 1986 Jan 29;134(2):624-31 PMID: 3080993
  13. Novel method for detection of beta-lactamases by using a chromogenic cephalosporin substrate.
    Antimicrob Agents Chemother. 1972 Apr;1(4):283-8 PMID: 4208895
  14. DNA sequencing with chain-terminating inhibitors.
    Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463-7 PMID: 271968
  15. Transfer of proteins across membranes. I. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane-bound ribosomes of murine myeloma.
    J Cell Biol. 1975 Dec;67(3):835-51 PMID: 811671
  16. Site-specific alteration of cysteine 176 and cysteine 234 in the lactose carrier of Escherichia coli.
    J Biol Chem. 1986 Sep 5;261(25):11765-9 PMID: 3528146
  17. Transfer of proteins across membranes.
    Annu Rev Biochem. 1981;50:317-48 PMID: 7023361
  18. Biosynthesis and periplasmic segregation of human proinsulin in Escherichia coli.
    Proc Natl Acad Sci U S A. 1981 Sep;78(9):5401-5 PMID: 7029534
  19. Secretion of Escherichia coli beta-lactamase from Bacillus subtilis by the aid of alpha-amylase signal sequence.
    Proc Natl Acad Sci U S A. 1982 Sep;79(18):5582-6 PMID: 6182566
  20. Method for blot-hybridization analysis of mRNA molecules from Bacillus subtilis.
    Gene. 1987;51(2-3):281-6 PMID: 2439407
  21. Prediction of protein antigenic determinants from amino acid sequences.
    Proc Natl Acad Sci U S A. 1981 Jun;78(6):3824-8 PMID: 6167991
  22. High frequency transformation of Bacillus subtilis protoplasts by plasmid DNA.
    Mol Gen Genet. 1979 Jan 5;168(1):111-5 PMID: 107388
  23. Mutations which alter the function of the signal sequence of the maltose binding protein of Escherichia coli.
    Nature. 1980 May 8;285(5760):78-81 PMID: 6990274
  24. Length and structural effect of signal peptides derived from Bacillus subtilis alpha-amylase on secretion of Escherichia coli beta-lactamase in B. subtilis cells.
    Nucleic Acids Res. 1984 Jul 11;12(13):5307-19 PMID: 6087281
  25. Prediction of the secondary structure of proteins from their amino acid sequence.
    Adv Enzymol Relat Areas Mol Biol. 1978;47:45-148 PMID: 364941
  26. A putative signal peptidase recognition site and sequence in eukaryotic and prokaryotic signal peptides.
    J Mol Biol. 1983 Jun 25;167(2):391-409 PMID: 6345794
  27. Importance of secondary structure in the signal sequence for protein secretion.
    Proc Natl Acad Sci U S A. 1983 Aug;80(15):4599-603 PMID: 6224220
  28. Secretion and membrane localization of proteins in Escherichia coli.
    CRC Crit Rev Biochem. 1980;7(4):339-71 PMID: 6993100
  29. Renaturation of enzymes after polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate.
    J Biol Chem. 1980 Aug 10;255(15):7467-73 PMID: 6156169
Article Info
Journal
Molecular & general genetics : MGG
Abbr.
Mol Gen Genet
ISSN
0026-8925
DOI
10.1007/BF00332223
Published
1989-03-00
Pages
1-9
Language
English
Region
Germany
NLM ID
0125036
Subset
IM
External Links
PubMed source DOI: 10.1007/BF00332223
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: [email protected]

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2