MilliporeSigma
  • Home
  • Search Results
  • Properties of PASP: a Pseudomonas protease capable of mediating corneal erosions.

Properties of PASP: a Pseudomonas protease capable of mediating corneal erosions.

Investigative ophthalmology & visual science (2009-03-04)
Aihua Tang, Mary E Marquart, Jonathan D Fratkin, Clare C McCormick, Armando R Caballero, Hattie P Gatlin, Richard J O'Callaghan
ABSTRACT

To analyze PASP in terms of its gene distribution and expression, its corneal pathologic effects, its enzymatic properties, and the protectiveness of the immune response to this protease. Twenty-five strains of P. aeruginosa were analyzed for the PASP gene and secreted protein by PCR and Western blot analysis, respectively. Active recombinant (r)PASP (10 microg/20 microL) or heat-inactivated rPASP was intrastromally injected into rabbit corneas. Pathologic changes were monitored by slit lamp examination (SLE) and histopathology. Purified rPASP was assayed for cleavage of collagens and susceptibility to TLCK. Rabbit antibody to rPASP was produced and tested for enzyme inactivation, and actively immunized rabbits were challenged by intrastromal injection of active rPASP (5 microg). All 25 strains of P. aeruginosa analyzed were positive for the PASP gene and protein. SLE scores of eyes injected with active rPASP were significantly higher than control eyes at all postinjection times (PI; P <or= 0.004). Histopathologic studies documented the destruction of the corneal epithelial layer and portions of the corneal stroma at 9 hours PI, and polymorphonuclear (PMN) leukocyte infiltration into the cornea by 24 hours after active rPASP injection. PASP cleaved type I and IV collagens and was susceptible to TLCK inhibition. PASP was present in the cytoplasm and periplasm, but only secreted PASP was enzymatically active. A high antibody titer (ELISA titer >or= 10,000) was produced, but this antibody did not protect against active rPASP challenge. PASP is a commonly produced Pseudomonas protease that can cleave collagens and cause corneal erosions.

MATERIALS
Product Number
Brand
Product Description

Millipore
Protein G–Agarose, lyophilized powder, Contains lactose stabilizers that must be removed prior to use.
Sigma-Aldrich
N-(p-Tosyl)-Gly-Pro-Lys 4-nitroanilide acetate salt, plasmin substrate
Millipore
Protein A–Agarose, lyophilized powder
Millipore
Protein A–Agarose, saline suspension
Millipore
Protein A–Agarose, lyophilized powder