title: Biosynthetic approach for functional protein microarrays.
abstract: Protein microarrays have emerged as an indispensable research tool for providing information about protein functions and interactions through high-throughput screening. Traditional methods for immobilizing biomolecules onto solid surfaces have been based on covalent and noncovalent binding, entrapment in semipermeable membranes, microencapsulation, sol gel, and hydrogel methods. Each of these techniques has its own strengths but fails to combine the most important tenets of a functional protein microarray such as covalent attachment, native protein conformation, homogeneity of the protein monolayer, control over active site orientation, and retention of protein activity. Here we present a selective and site-directed covalent immobilization technique for proteins via a benzoxazine ring formation through a Diels-Alder reaction in water and a genetically encoded 3-amino-L-tyrosine (3-NH(2)Tyr) amino acid. Fully functional protein microarrays, with monolayer arrangements and complete control over their orientations, were generated using this strategy.
authors.affiliation: Department of Chemistry and Biochemistry, University of Texas at Arlington, Arlington, TX 76019, USA.
authors.forename: Brian
authors.initials: B
authors.lastname: Stamos
authors.forename: Leticia
authors.initials: L
authors.lastname: Loredo
authors.forename: Subhash
authors.initials: S
authors.lastname: Chand
authors.forename: Tuan V
authors.initials: TV
authors.lastname: Phan
authors.forename: Yanbo
authors.initials: Y
authors.lastname: Zhang
authors.forename: Sridev
authors.initials: S
authors.lastname: Mohapatra
authors.forename: Krishnan
authors.initials: K
authors.lastname: Rajeshwar
authors.forename: Roshan
authors.initials: R
authors.lastname: Perera
mesh_terms: D048588:Benzoxazines / Q000737:chemistry; D004926:Escherichia coli; D057166:High-Throughput Screening Assays; D055767:Immobilized Proteins / Q000737:chemistry* / Q000235:genetics; D018625:Microscopy, Atomic Force; D010504:Periodic Acid / Q000737:chemistry; D010957:Plasmids; D011092:Polyethylene Glycols / Q000737:chemistry; D040081:Protein Array Analysis / Q000379:methods*; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D012822:Silicon Dioxide / Q000737:chemistry; D012996:Solutions; D021241:Spectrometry, Mass, Electrospray Ionization; D013499:Surface Properties; D014443:Tyrosine / Q000031:analogs & derivatives / Q000737:chemistry; D014867:Water
chemical_list: D048588:Benzoxazines; D055767:Immobilized Proteins; D011506:Proteins; D011994:Recombinant Proteins; D012996:Solutions; D014867:Water; D010504:Periodic Acid; C019253:3-aminotyrosine; D011092:Polyethylene Glycols; D014443:Tyrosine; D012822:Silicon Dioxide; C009288:metaperiodate