System to Produce Biotinylated Proteins
Any protein, antibody, or antibody fragments can be biotinylated post-translationally Shortens and streamlines the current process of protein biotinylation by eliminating the need to purify and treat secreted proteins with exogenous reagents Biotinylation efficiency is >90% Reduces chance of protein degradation
Produce uniformly biotinylated secreted proteins for in vivo by both versions of biotin ligase, secreted and ER-retained May be used for streptavidin/avidin-biotin technology Stable cell line expressing ER-retained BirA (biotin protein ligase of E. Coli) can be generated for research purposes
UCLA researchers have developed a system that allows metabolic biotinylation of secreted proteins in vivo by both versions of biotin ligase (secreted and ER-retained). The system is a general approach for production of site-specific biotinylated proteins for streptavidin/avidin-biotin technology. The biotinylation technology also shortens and streamlines the lengthy process of traditional methods and reduces the chance of protein degradation during in vitro biotinylation by exogenous ligase. Not only is this system a significant improvement over traditional chemical methods, the system produces uniformly biotinylated proteins without altering biological functions or binding specificity. Researchers also generated a stable cell line expressing ER-retained BirA, the biotin protein ligase of E. Coli
Patent Number: US8043830B2
Application Number: US2009363678A
Inventor: Barat, Bhaswati | Wu, Anna M.
Priority Date: 1 Feb 2008
Priority Number: US8043830B2
Application Date: 30 Jan 2009
Publication Date: 25 Oct 2011
IPC Current: C12P002100 | C12P002104
US Class: 4350691 | 4350697
Assignee Applicant: The Regents of the University of California
Title: Biotin-ligase system for secretion of biotinylated protein
Usefulness: Biotin-ligase system for secretion of biotinylated protein
Summary: The method, eukaryotic cell line, and kit are useful for producing a biotinylated polypeptide, and a secreted heterologous biotinylated polypeptide (all claimed).
Novelty: Producing a biotinylated polypeptide, by expressing a heterologous target polypeptide with a biotinylation acceptor sequence in a eukaryotic cell, expressing in the same eukaryotic cell a heterologous biotin protein ligase, and purifying
生物醫學
DNA /基因工程
8043830
State of Development Investigators have tested the system to biotinylated proteins in vivo by biotin ligase. Background Biotin (vitamin H) is an essential coenzyme that is also used to tag proteins for detection, labeling, and purification purposes. The process of adding biotin to proteins is called biotinylation. Biotin labeling has also been applied to drug targeting and viral gene therapy vector-targeting strategies. Traditionally, biotin labeling has been performed in vitro by chemical methods. The problem with these chemical methods is that the random and heterogeneous modifications can lead to the inactivation of biological function after mixing with streptavidin or avidin. Antibody biotinylation especially leads to heterogeneous conjugates. Therefore, there is a need for a method that will uniformly biotinylated proteins without altering binding properties and resulting in loss of affinity. Related Materials Additional Technologies by these Inventors Tech ID/UC Case 20232/2008-501-0 Related Cases 2008-501-0
美國
