Biomaterial Database

Site-specific, covalent attachment of poly(dT)-modified peptides to solid surfaces for microarrays. 2007

Naoki Kimura, and Takashi Okegawa, and Kiyokazu Yamazaki, and Koji Matsuoka

Research and Development Center, Nisshinbo Industries Inc, Chiba, Japan. n-kimu@nisshinbo.co.jp

The present study reported proof-of-principle for a kinase assay approach that can detect specific peptide phosphorylation events. The method involves attachment of peptides onto commercial aminosilane and polycarbodiimide-coated glass slides, using a newly developed DNattach linker system that consists of a poly(dT) tail (Nisshinbo Industries Inc.), followed by a detection step using fluorescently labeled antiphosphoamino acid antibodies. The linker-modified peptides are efficiently synthesized by Michael addition between maleimido-modified peptides and thiol-containing DNattach. Specific covalent immobilization of the modified peptides onto aminosilane and poly carbodiimide-coated slides is then achieved by short exposure to UV-light. Highly selective and quantitative recognition by standard antiphosphoamino acid antibodies (antiphosphotyrosine and anti-phosphoGFAP) and kinases (c-Src and PKA) to the corresponding modified peptides on the microarray spots is demonstrated. Furthermore, we found that this immobilization method provides greater signal-to-noise ratio and better discrimination ability of phosphorylated amino acids than does the conventional immobilization technique. The phosphorylation pattern of target sequences, detected using fluorescently labeled antiphosphoamino acid antibodies, revealed that the linker system preference of the kinase is determined by its activity profile.

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D010455	Peptides	Members of the class of compounds composed of AMINO ACIDS joined together by peptide bonds between adjacent amino acids into linear, branched or cyclical structures. OLIGOPEPTIDES are composed of approximately 2-12 amino acids. Polypeptides are composed of approximately 13 or more amino acids. PROTEINS are considered to be larger versions of peptides that can form into complex structures such as ENZYMES and RECEPTORS.	Peptide,Polypeptide,Polypeptides
D010766	Phosphorylation	The introduction of a phosphoryl group into a compound through the formation of an ester bond between the compound and a phosphorus moiety.	Phosphorylations
D011071	Poly T	A group of thymine nucleotides in which the phosphate residues of each thymine nucleotide act as bridges in forming diester linkages between the deoxyribose moieties.	Poly dT,Polythymidylic Acids,Thymine Polynucleotides,Polydeoxythymidylate,Acids, Polythymidylic,Polynucleotides, Thymine,dT, Poly
D011505	Protein-Tyrosine Kinases	Protein kinases that catalyze the PHOSPHORYLATION of TYROSINE residues in proteins with ATP or other nucleotides as phosphate donors.	Tyrosine Protein Kinase,Tyrosine-Specific Protein Kinase,Protein-Tyrosine Kinase,Tyrosine Kinase,Tyrosine Protein Kinases,Tyrosine-Specific Protein Kinases,Tyrosylprotein Kinase,Kinase, Protein-Tyrosine,Kinase, Tyrosine,Kinase, Tyrosine Protein,Kinase, Tyrosine-Specific Protein,Kinase, Tyrosylprotein,Kinases, Protein-Tyrosine,Kinases, Tyrosine Protein,Kinases, Tyrosine-Specific Protein,Protein Kinase, Tyrosine-Specific,Protein Kinases, Tyrosine,Protein Kinases, Tyrosine-Specific,Protein Tyrosine Kinase,Protein Tyrosine Kinases,Tyrosine Specific Protein Kinase,Tyrosine Specific Protein Kinases
D002234	Carbodiimides	Compounds with the general formula RN	Carbodiimide
D005898	Glass	Hard, amorphous, brittle, inorganic, usually transparent, polymerous silicate of basic oxides, usually potassium or sodium. It is used in the form of hard sheets, vessels, tubing, fibers, ceramics, beads, etc.
D000081247	CSK Tyrosine-Protein Kinase	Protein tyrosine kinases that phosphorylate tyrosine residues located in the C-terminal tails of SRC-FAMILY KINASES.	C-Terminal Src Kinase,Carboxy-Terminal Src Kinase,Protein Tyrosine Kinase p50(csk),Protein-Tyrosine Kinase C-Terminal Src Kinase,Protein-Tyrosine Kinase c-src,Tyrosine Protein Kinase p50csk,Tyrosine-Protein Kinase CSK,c-src Kinase,c-src Tyrosine Kinase,CSK-src,p50(csk),C Terminal Src Kinase,CSK Tyrosine Protein Kinase,CSK src,CSK, Tyrosine-Protein Kinase,Carboxy Terminal Src Kinase,Kinase CSK, Tyrosine-Protein,Kinase c-src, Protein-Tyrosine,Kinase, C-Terminal Src,Kinase, CSK Tyrosine-Protein,Kinase, Carboxy-Terminal Src,Kinase, c-src,Kinase, c-src Tyrosine,Protein Tyrosine Kinase C Terminal Src Kinase,Protein Tyrosine Kinase c src,Src Kinase, C-Terminal,Src Kinase, Carboxy-Terminal,Tyrosine Kinase, c-src,Tyrosine Protein Kinase CSK,Tyrosine-Protein Kinase, CSK,c src Kinase,c src Tyrosine Kinase,c-src, Protein-Tyrosine Kinase
D000586	Amination	The creation of an amine. It can be produced by the addition of an amino group to an organic compound or reduction of a nitro group.	Aminations
D012680	Sensitivity and Specificity	Binary classification measures to assess test results. Sensitivity or recall rate is the proportion of true positives. Specificity is the probability of correctly determining the absence of a condition. (From Last, Dictionary of Epidemiology, 2d ed)	Specificity,Sensitivity,Specificity and Sensitivity