Protein microarray on-demand: a novel protein microarray system.
We describe a novel, simple and low-cost protein microarray strategy wherein the microarrays are generated by printing expression ready plasmid DNAs onto slides that can be converted into protein arrays on-demand. The printed expression plasmids serve dual purposes as they not only direct the synthesis of the protein of interest; they also serve to capture the newly synthesized proteins through a high affinity DNA-protein interaction. To accomplish this we have exploited the high-affinity binding (approximately 3-7 x 10 (-13) M) of E. coli Tus protein to Ter, a 20 bp DNA sequence involved in the regulation of E. coli DNA replication. In our system, each protein of interest is synthesized as a Tus fusion protein and each expression construct directing the protein synthesis contains embedded Ter DNA sequence. The embedded Ter sequence functions as a capture reagent for the newly synthesized Tus fusion protein. This "all DNA" microarray can be converted to a protein microarray on-demand without need for any additional capture reagent.
Chatterjee
Deb K
DK
Protein Expression Laboratory, SAIC-Frederick, Inc., NCI-Frederick, Frederick, Maryland, USA. chatterjee@mail.ncifcrf.gov
Sitaraman
Kalavathy
K
Baptista
Cassio
C
Hartley
James
J
Hill
Thomas M
TM
Munroe
David J
DJ
D001426:Bacterial Proteins / Q000737:chemistry; D001483:Base Sequence; D019295:Computational Biology; D004261:DNA Replication; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010957:Plasmids / Q000378:metabolism; D040081:Protein Array Analysis / Q000295:instrumentation* / Q000379:methods*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D011993:Recombinant Fusion Proteins / Q000737:chemistry
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins