Download Centennial Honors College Western Illinois University Undergraduate Research Day 2014

Centennial Honors College
Western Illinois University
Undergraduate Research Day 2014
Poster Presentation
Evaluation of New Cathepsin K Inhibitors
Amanda Ellison and David VanDerway
Faculty Mentors: Lisa Wen and Jenq-Kuen Huang
Chemistry
Cathepsin K is an enzyme that is involved with collagenolytic activity, bone resorption,
and bone degradation. It has been found at a higher level in people with rheumatoid
arthritis, prostate cancer and even breast cancer due to its role in matrix homeostasis.
Due to its involvement in these detrimental conditions and medical importance, we hope
to aid in the development of drugs that will inhibit this enzyme by producing a
substantial amount of its active form that can be tested and subjected to potential
inhibitors for drug screening. The recombinant human procathepsin K has been
overexpressed in E. coli, purified from inclusion bodies (insoluble protein aggregates).
The inactive Procathepsin K is then treated with pepsin that converts the enzyme to its
active form, cathepsin K. The success of activation is monitored by SDS-polyacrylamide
gel electrophoresis (SDS-PAGE) on 15% acrylamide slab gels, which separates protein
by size. Activated cathepsin K is smaller than procathepsin K and should migrate faster
in an SDS-PAGE. Cathepsin K inhibition assay is performed by monitoring changes in
absorption at 405 nm spectrophotometrically in the presence of N-Carbobenzoxy-LPhenylalanyl-Arginine-4-nitroanilide hydrochloride as the chromogenic substrate. Then
an IC-50 analysis is performed to see what exact concentration of the inhibitor is
needed for the inhibition of 50% of the Cathepsin K enzyme. Data analysis of the
effectiveness of these inhibitors will be used to foster future development of newer
effective inhibitors for the treatment osteoporosis and possibly rheumatoid arthritis and
cancer. This research will be completed in the spring of 2015.