What question did this study set out to answer?

To determine the genetic relationship between the 49-kDa and 53-kDa forms of exoenzyme S from Pseudomonas aeruginosa and test whether the smaller protein arises from carboxy-terminal cleavage of the larger form.

March 1, 1996Journal of BacteriologyOpen Access

Genetic relationship between the 53- and 49-kilodalton forms of exoenzyme S from Pseudomonas aeruginosa

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Authors

TYTimothy L. YahrUniversity of Iowa JBJoseph BarbieriMedical College of Wisconsin DFDara W. FrankMedical College of Wisconsin

Discussion

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Implication

Genetic study demonstrates that distinct genes encode the 53-kDa and 49-kDa exoenzyme S forms in Pseudomonas aeruginosa, indicating they are separate toxins rather than cleavage products.

Key Points

To determine the genetic relationship between the 49-kDa and 53-kDa forms of exoenzyme S from Pseudomonas aeruginosa and test whether the smaller protein arises from carboxy-terminal cleavage of the larger form.
Screened chromosomal digests and a genomic library from an exoS deletion mutant of Pseudomonas aeruginosa (388deltaexoS::TC) via Southern blot using an internal exoS probe.
Cloned the identified open reading frame, exoT, into a T7 expression system in Escherichia coli to generate recombinant 53-kDa protein (Exo53).
Assayed ADP-ribosyltransferase activity of histidine-tagged Exo53 and generated an allelic-replacement knockout of exoT in Pseudomonas aeruginosa.
Identified exoT, an open reading frame encoding a 457-amino-acid protein with 75% identity to ExoS that produces a 53-kDa protein cross-reacting with anti-exoenzyme S antibodies.
Observed that recombinant Exo53 possesses approximately 0.2% of the ADP-ribosyltransferase activity of recombinant ExoS.
Demonstrated that targeted inactivation of exoT yields an Exo53-deficient phenotype without altering ExoS expression, confirming that separate genes encode the two forms.

Cite This Study

Yahr et al. (1996) studied this question.

synapsesocial.com/papers/6a72c44d35aa2c282ce36d6f https://doi.org/10.1128/jb.178.5.1412-1419.1996

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March 1, 1996Journal of BacteriologyOpen Access

Genetic relationship between the 53- and 49-kilodalton forms of exoenzyme S from Pseudomonas aeruginosa

View Full Paper

Ask AI

Bookmark

Authors

TYTimothy L. YahrUniversity of Iowa JBJoseph BarbieriMedical College of Wisconsin DFDara W. FrankMedical College of Wisconsin

Discussion

Member takes

Implication

Genetic study demonstrates that distinct genes encode the 53-kDa and 49-kDa exoenzyme S forms in Pseudomonas aeruginosa, indicating they are separate toxins rather than cleavage products.

Key Points

To determine the genetic relationship between the 49-kDa and 53-kDa forms of exoenzyme S from Pseudomonas aeruginosa and test whether the smaller protein arises from carboxy-terminal cleavage of the larger form.
Screened chromosomal digests and a genomic library from an exoS deletion mutant of Pseudomonas aeruginosa (388deltaexoS::TC) via Southern blot using an internal exoS probe.
Cloned the identified open reading frame, exoT, into a T7 expression system in Escherichia coli to generate recombinant 53-kDa protein (Exo53).
Assayed ADP-ribosyltransferase activity of histidine-tagged Exo53 and generated an allelic-replacement knockout of exoT in Pseudomonas aeruginosa.
Identified exoT, an open reading frame encoding a 457-amino-acid protein with 75% identity to ExoS that produces a 53-kDa protein cross-reacting with anti-exoenzyme S antibodies.
Observed that recombinant Exo53 possesses approximately 0.2% of the ADP-ribosyltransferase activity of recombinant ExoS.
Demonstrated that targeted inactivation of exoT yields an Exo53-deficient phenotype without altering ExoS expression, confirming that separate genes encode the two forms.

Cite This Study

Yahr et al. (1996) studied this question.

synapsesocial.com/papers/6a72c44d35aa2c282ce36d6f https://doi.org/10.1128/jb.178.5.1412-1419.1996

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