Purification and characterization of a serine protease and chitinases from Paecilomyces lilacinus and detection of chitinase activity on 2D gels

Alamgir Khan*, Keith Williams, Mark P. Molloy, Helena Nevalainen

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    73 Citations (Scopus)

    Abstract

    The filamentous fungus Paecilomyces lilacinus is currently developed as a biocontrol agent against plant parasitic nematodes. Nematode eggs and cuticles are the infection sites for biocontrol agents that penetrate by the production of lytic enzymes. P. lilacinus was cultured in liquid media and proteases and chitinases were induced by the introduction of egg yolk and chitin, respectively. A serine protease was purified from a culture medium using Sepharose-bacitracin affinity column. The protease occurred in three forms, two of which were C-terminally truncated. Chitinase activity was also observed in the culture supernatant, and after separation by isoelectric focusing six proteins were detected that showed activity. Chitinase activity was further confirmed on non-denaturing one-dimensional (1D) and two-dimensional (2D) gels using a sandwich assay with glycol chitin as a substrate. Two of the proteins had similarities with endochitinases as shown by their N-terminal amino acid sequences.

    Original languageEnglish
    Pages (from-to)210-220
    Number of pages11
    JournalProtein Expression and Purification
    Volume32
    Issue number2
    DOIs
    Publication statusPublished - Dec 2003

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