Cloning the structural gene for the 49-kDa form of exoenzyme S (exoS) from Pseudomonas aeruginosa strain 388

Scott M Kulich; Timothy L Yahr; Liane M Mende-Mueller; Joseph T Barbieri; Dara W Frank

doi:10.1016/S0021-9258(17)34078-4

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Cloning the structural gene for the 49-kDa form of exoenzyme S (exoS) from Pseudomonas aeruginosa strain 388

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Cloning the structural gene for the 49-kDa form of exoenzyme S (exoS) from Pseudomonas aeruginosa strain 388

Scott M Kulich, Timothy L Yahr, Liane M Mende-Mueller, Joseph T Barbieri and Dara W Frank

The Journal of biological chemistry, Vol.269(14), pp.10431-10437

04/08/1994

DOI: 10.1016/S0021-9258(17)34078-4

PMID: 8144626

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Abstract

We report the purification and proteolytic characterization of the 49-kDa form of exoenzyme S and the cloning of the structural gene for the 49-kDa form of exoenzyme S (exoS). The 49-kDa form of exoenzyme S was purified from SDS-polyacrylamide gels. Conditions were established that allowed efficient trypsin digestion of the 49-kDa form of exoenzyme S. Amino acid sequence determination of the amino terminus and tryptic peptides of the 49-kDa form of exoenzyme S allowed the generation of degenerate oligonucleotides, which were used to amplify DNA encoding an amino-terminal sequence and an internal sequence of the 49-kDa form of exoenzyme S. These DNA fragments were used to clone the entire structural gene for the 49-kDa form of exoenzyme S (exoS) from a cosmid library of Pseudomonas aeruginosa strain 388. The 49-kDa form of exoenzyme S (ExoS) is predicted to be a 453 amino acid protein. The predicted amino acid sequence indicates that ExoS is secreted from Pseudomonas without cleavage of an amino-terminal sequence. BESTFIT analysis identified three regions of alignment between ExoS and the active site of Escherichia coli heat-labile enterotoxin. One region of homology appears to be shared among several members of the family of bacterial ADP-ribosyltransferases.

Amino Acid Sequence

Genes, Bacterial

Oligonucleotide Probes

Electrophoresis, Polyacrylamide Gel

Molecular Sequence Data

Chromatography, High Pressure Liquid

Chromosomes, Bacterial

Restriction Mapping

DNA, Bacterial

Hydrolysis

Sequence Homology, Amino Acid

Poly(ADP-ribose) Polymerases - metabolism

Trypsin

Poly(ADP-ribose) Polymerases - genetics

Pseudomonas aeruginosa - genetics

Bacterial Toxins

Pseudomonas aeruginosa - enzymology

ADP Ribose Transferases

Base Sequence

Cloning, Molecular

Poly(ADP-ribose) Polymerases - isolation & purification

Details

Title: Subtitle: Cloning the structural gene for the 49-kDa form of exoenzyme S (exoS) from Pseudomonas aeruginosa strain 388
Creators: Scott M Kulich - Department of Microbiology, Medical College of Wisconsin, Milwaukee 53226
Timothy L Yahr
Liane M Mende-Mueller
Joseph T Barbieri
Dara W Frank
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.269(14), pp.10431-10437
DOI: 10.1016/S0021-9258(17)34078-4
PMID: 8144626
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Publisher: United States
Grant note: AI01087 / NIAID NIH HHS AI31665 / NIAID NIH HHS
Language: English
Date published: 04/08/1994
Academic Unit: Microbiology and Immunology
Record Identifier: 9984001107702771

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