Targeting bacterial virulence as a novel antibiotic and vaccine approach

Where a bacterial virulence factor such as SpyCEP is alone, sufficient to cause disease, that virulence factor represents a suitable target for both vaccine development and therapeutic approaches.Aim 1 Detailed characterization of the enzymatic activity of purified recombinant SpyCEP
SpyCEP is known to cleave and inactivate CXCL8 and other ELR+ chemokines resulting in impaired neutrophil recruitment to the site of infection. It is likely that cleavage also impacts many other known CXCL8-mediated effects. Due to a prior lack of adequate quantities of purified enzyme, a systematic study of enzyme kinetics has not been previously undertaken, though would be required for development of any inhibitor or assay of inhibition.
As such, at the start of this work I will undertake kinetic studies using purified SpyCEP enzyme from GAS supernatant and recombinantly produced enzyme (all available in the laboratory) in order to determine the KM and Vmax parameters of SpyCEP using ELISA to monitor CXCL8 cleavage and SDS-PAGE or MS-based analyses if necessary.

SpyCEP appears to have a number of additional activities that may relate to previously unknown effects of CXCL8, or to additional previously unknown substrates. These should be clarified in order to more systematically assess the efficacy of any vaccine or therapeutic.