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 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 degrees C (the temperature of the flea vector) to 37 degrees C (the temperature of the mammalian host). To model this process, Y. pestis cells were grown at 26 degrees C and 37 degrees C in a Ca(2+)-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 degrees C. Two of the proteins predominately expressed at 37 degrees C 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.
Ciphergen Biosystems, Fremont, CA, USA. vanitha@ciphergen.com
V
V
Thulasiraman
S L
SL
McCutchen-Maloney
V L
VL
Motin
E
E
Garcia
D001426:Bacterial Proteins / Q000032:analysis*; D007834:Lasers; D008970:Molecular Weight; D014774:Virulence; D015010:Yersinia pestis / Q000737:chemistry / Q000472:pathogenicity*
D001426:Bacterial Proteins