Specific Labeling of Protein with Zinc Finger Tags and Use of Zinc-Finger-Tagged Proteins for Analysis
AssigneeThe Scripps Research Institute
InventorCarlos Barbas, III
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Abstract
Fusion proteins including zinc finger tags that bind in a sequence-specific manner and a peptide, polypeptide, or protein can be prepared and expressed. Such fusion proteins can be used to generate protein arrays by binding the zinc finger tags to a DNA array. The fusion proteins can also be used to label cell surfaces with DNA tags. Fusion proteins according to the invention can be used to localize the peptide, polypeptide, or protein that is incorporated into the fusion protein by using a labeled DNA, such as a fluorescent DNA. The invention further includes vectors and host cells, as well as a method of analyzing double-stranded DNA.
Claims82
An array comprising:
- (a) a solid support;
- (b) a plurality of nucleotide sequences attached to the solid support;
- and
- (c) a plurality of fusion proteins specifically and noncovalently bound to the plurality of nucleotide sequences, each fusion protein comprising:
- (1) a protein, peptide, or polypeptide of interest;
- and (2) a zinc finger protein tag, wherein each zinc finger protein tag has specific binding affinity for only one of the nucleotide sequences attached to the solid support.
A fusion protein comprising:
- (a) a protein, polypeptide, or peptide of interest;
- and
- (b) at least one zinc finger tag in a single polypeptide;
- such that the protein, polypeptide, or protein of interest substantially maintains its three-dimensional conformation and activity, and the zinc finger tag substantially maintains its sequence-specific nucleotide sequence binding activity.
A method for labeling the cell membrane of a cell comprising the steps of:
- (a) transforming or transfecting a host cell with a nucleic acid sequence that encodes a fusion protein that is a fusion of a membrane protein with a zinc finger tag such that the cell expresses the fusion protein;
- (b) culturing the transformed or transfected cell under conditions such that the fusion protein is expressed and is incorporated in the cell membrane of the cell;
- (c) contacting the cell expressing the fusion protein incorporated in the membrane with a labeled DNA molecule that binds the zinc finger tag of the fusion protein in a sequence-specific manner;
- and
- (d) detecting the label of the labeled DNA molecule on the cell surface.
Dependencies are inferred from the claim text.