Detection and identification of virulence factors in Yersinia pestis using SELDI ProteinChip® system

A rapid method for the detection, purification, and identification of proteins in bacterial extracts was developed using surface enhanced laser desorption/ionization (SELDI) ProteinChip(R) technology. The effectiveness of this technique for monitoring the expression and identification of temperature- and calcium-regulated virulence factors of Yersinia pestis, the bacterium that causes human plague, is demonstrated. Y. pestis infection of its mammalian host is thought to be accompanied by rapid up-regulation of a number of genes following a shift from 26 degreesC (the temperature of the flea vector) to 37 degreesC (the temperature of the mammalian host). To model this process, Y. pestis cells were grown at 26 degreesC and 37 degreesC in a Ca2+-deficient medium. through an initial protein profiling of the crude bacterial extract on strong anion exchange and copper affinity, ProteinChip arrays detected five proteins that were up-regulated and three proteins that were down-regulated at 37 degreesC. Two of the proteins predominately expressed at 37 degreesC were semi-purified in less than two days. The two proteins were identified as catalase-peroxidase and Antigen 4. Aside from its speed, a salient feature of the SELDI technique is the microgram amounts of crude sample required for analysis.