Microarray of DNA-protein complexes on poly-3-hydroxybutyrate surface for pathogen detection

作者：Park Tae Jung; Yoo Seung Min; Keum Ki Chang; Lee Sang Yup^*

来源：Analytical and Bioanalytical Chemistry, 2009, 393(6-7): 1639-1647.

DOI：10.1007/s00216-008-2574-y

• Protein microarray
• DNA chip
• Poly-3-hydroxybutyrate
• Pathogen detection
• PHB depolymerase
• Substrate-binding domain

A novel strategy was developed for the specific immobilization of DNA probes on poly-3-hydroxybutyrate (PHB) surface by using the substrate-binding domain (SBD) of PHB depolymerase as an active binding motif. To demonstrate whether this method can be used for the detection of clinical pathogens, the pathogen-specific biotin-labeled DNA probes were immobilized via core streptavidin (cSA) fused to the SBD. The pathogen-specific 15-mer oligonucleotide probes were designed for four model pathogens, while the target DNAs were prepared by PCR using universal primers. The complex of pathogen-specific probes and cSA-SBD fusion protein was immobilized on the PHB-coated slide by microspotting. This DNA-protein complex microarray was able to successfully diagnose Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa. Furthermore, the specific pathogens could be diagnosed in the presence of other microorganisms. Thus, the DNA-protein complex microarray platform technology employing PHB and the SBD reported here can be widely used for the detection of DNA-DNA and DNA-biomolecule interactions without synthetic or chemical modification of biomolecules or solid surface.

• 出版日期2009-3