Abstract:
Candida cell wall proteomics is a challenging area of research. Extraction of cell wall proteins needs to
be done carefully to obtain reproducible results. We used two different methods for extraction of
candida cell wall proteins and documented the modifications required. In the course of extraction, we
also evaluated efficacy of cell wall disruption by mechanical procedure of homogenization with glass
beads to increase the protein yield and purity. For extraction of candida cell wall proteins, we utilized
detergents like Sodium dodecyl sulphate (SDS), dithiothreitol (DTT), tris base and chitinase. This
method was further optimized using homogenization of candida cells with glass beads over different
time intervals. Amount of cell disruption at each time interval of bead beating using two varieties of
beads was evaluated by gram stain, culture, protein estimation and SDS PAGE subsequently.
We observed that method using detergents and Chitinase yielded better quality proteins, less
contaminated with salts. For homogenization, bead beating with glass beads of 0.5 mm diameter over
18 minutes was found to be superior. Our initial experiments revealed limits of both the methods in
candida cell wall protein extraction. Mechanical disruption of the cells could be further optimized using
routinely available vortex mixer to increase final protein yield and gave consistent and reproducible
results.
