TurboID is a proximity-dependent biotin identification (BioID) enzyme derived from the bacterial biotin ligase (BirA). It was developed by Jonathan S. Weissman and Alice Y. Ting in 2018 as an enhanced version of the original BioID method. TurboID allows researchers to investigate protein-protein interactions and spatial proximities with improved sensitivity, temporal control, and labeling efficiency.
The principle underlying TurboID is based on proximity-dependent biotinylation. In TurboID, the protein of interest is fused to the TurboID enzyme, which has a mutated biotin ligase domain. When the TurboID fusion protein is expressed in cells or organisms and supplied with biotin, the enzyme can biotinylate nearby proteins within a certain radius. Biotinylation occurs when the enzyme transfers biotin from biotin-AMP to the lysine residues of proximal proteins.
Proximity-dependent biotinylation catalyzed by TurboID and split-TurboID. (Cho, 2020)
The workflow of TurboID typically involves the following steps:
Constructing TurboID Fusion Protein: The gene encoding the protein of interest is genetically fused to the TurboID enzyme coding sequence using molecular cloning techniques. This fusion construct is then introduced into the target cells or organisms.
Expression and Biotinylation: After successful transfection or transduction, the TurboID fusion protein is expressed in the cellular system of interest. Biotin, either exogenously supplied or endogenously produced, is then introduced into the system. TurboID biotinylates proximal proteins in the presence of biotin and ATP.
Isolation of Biotinylated Proteins: Biotinylated proteins are captured using streptavidin affinity purification. The biotinylated proteins can be specifically enriched by binding to streptavidin beads or other streptavidin-based matrices.
Protein Identification and Analysis: The enriched biotinylated proteins are eluted from the streptavidin beads and subjected to mass spectrometry-based proteomic analysis. The identified proteins can then be further analyzed for interaction networks, functional annotations, and biological insights.
Mapping Protein-Protein Interactions: TurboID enables the systematic identification of protein-protein interaction networks. By fusing TurboID to a protein of interest, researchers can identify and characterize novel interacting partners, thereby unraveling complex cellular pathways and protein complexes.
Investigating Subcellular Localization: TurboID can be employed to determine the subcellular localization of proteins and understand their spatial relationships within cellular compartments. This information is crucial for elucidating organelle dynamics, membrane trafficking, and signaling processes.
Studying Protein Dynamics: TurboID facilitates the analysis of temporal changes in protein interactions. By controlling the duration of biotin labeling, researchers can investigate the dynamics of protein-protein interactions during different cellular processes, such as cell division, differentiation, and response to external stimuli.
Identifying Drug Targets: TurboID can aid in the discovery of potential drug targets by identifying proteins that interact with specific target proteins or protein complexes. This information can guide the development of therapeutic interventions and precision medicine approaches.
Creative Proteomics offers TurboID services to researchers worldwide. Our experienced team of experts utilizes state-of-the-art technologies and advanced bioinformatics tools to perform TurboID experiments and data analysis. Our TurboID service includes:
TurboID Construct Design: We assist researchers in designing TurboID fusion constructs tailored to their specific needs and experimental systems.
Expression and Biotinylation: Our team ensures efficient expression of TurboID fusion proteins and optimal biotinylation conditions to achieve high-quality labeling.
Protein Isolation and Enrichment: We employ robust streptavidin-based techniques to isolate and enrich biotinylated proteins, ensuring maximum capture efficiency and minimal background noise.
Mass Spectrometry-based Proteomic Analysis: Our advanced mass spectrometry platforms enable comprehensive and in-depth analysis of biotinylated proteins, facilitating the identification of interacting partners and functional annotations.
Data Interpretation and Reporting: We provide detailed data interpretation and comprehensive reports, including protein interaction networks, Gene Ontology analysis, and pathway enrichment analysis, to help researchers gain valuable insights from TurboID experiments.
With the TurboID service provided by Creative Proteomics, you can leverage this cutting-edge technology to unlock new insights into cellular processes. Contact us to learn more.