This control plasmid contains the gene encoding the Cytochrome C oxidase, subunit 8-2 (COX8-2) protein cloned upstream of the SNAPf coding sequence in pSNAPf, as a fusion to the N- terminus of the SNAP-tag. Cytochrome C oxidase is located in the inner mitochondrial membrane and is the terminal enzyme of the respiratory chain. The COX8-2-SNAPf fusion protein gives mitochondrial fluorescence when labeled with SNAP-Cell™ substrates. The full sequence and map for pSNAPf-Cox8A can be downloaded.
The SNAP-tag is a novel tool for protein research, allowing the specific, covalent attachment of virtually any molecule to a protein of interest. The SNAP-tag is a small protein based on human O6-alkylguanine-DNA-alkyltransferase (hAGT). SNAP-tag substrates are derivatives of benzyl purines and benzyl pyrimidines. In the labeling reaction, the substituted benzyl group of the substrate is covalently attached to the SNAP-tag.
pSNAPf-Cox8A contains an improved version of SNAP-tag, termed SNAPf. SNAPf displays faster kinetics in in vitro labeling and fast, specific and efficient labeling in live and fixed cell applications, thereby rendering it a desired research tool for analysis of protein dynamics.
There are two steps to using this system: sub cloning and expression of the protein of interest as a SNAPf fusion, and labeling of the fusion with the SNAP-tag substrate of choice. Expression of the SNAPf-Cox8A fusion protein is described in this document. The labeling of the fusion proteins with SNAP-tag substrates is described in the instructions supplied with SNAP-tag substrates.
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