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In vivo Biotinylation Based 

Method for the Study of Protein-

Protein Proximity in Eukaryotic 

Cells 

 

Arman Kulyyassov1, Erlan 

Ramanculov2, Vasily Ogryzko3 

 
1National Center for Biotechnology, 

Almaty, Kazakhstan; 2Interdisciplinary 

Instrumentation Center, Nazarbayev 

University, Astana, Kazakhstan; 3Institut 

Gustave Roussy, Villejuif, France 

 

Vol. 2, Suppl. (2013)   |   ISSN 2166-7403 (online)    

DOI 10.5195/cajgh.2013.96   |   http://cajgh.pitt.edu 

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KULYYASSOV 

 

 

This work is licensed under a Creative Commons Attribution 3.0 United States License. 

 

This journal is published by the University Library System of the University of Pittsburgh as part  

of its D-Scribe Digital Publishing Program and is cosponsored by the University of Pittsburgh Press. 

 

Central Asian Journal of Global Health 

Volume 2, Suppl. (2013)  |  ISSN 2166-7403 (online)  |  DOI 10.5195/cajgh.2013.96  |  http://cajgh.pitt.edu 

  

 

Abstract 

Introduction: The spatiotemporal order plays an important role in cell functioning and is affected in many pathologies such as 

cancer and neurodegenerative diseases. One of the ultimate goals of molecular biology is reconstruction of the spatiotemporal 

structure of a living cell at the molecular level. This task includes determination of proximities between different molecular 

components in the cell and monitoring their time- and physiological state-dependent changes. In many cases, proximity between 

macromolecules arises due to their interactions; however, the contribution of dynamic self-organization in generation of 

spatiotemporal order is emerging as another viable possibility. Specifically, in proteomics, this implies that the detection of 

protein-protein proximity is a more general task than gaining information about physical interactions between proteins, as it could 

detail aspects of spatial order in vivo that are challenging to reconstitute in binding experiments in vitro. 

Methods: In this work, we have developed a method of monitoring protein-protein proximity in vivo. For this purpose, the BirA 

was fused to one of the interaction partners, whereas the BAP was modified to make the detection of its biotinylation possible by 

mass spectrometry.  

Results: Using several experimental systems, we showed that the biotinylation is interaction dependent. In addition, we 

demonstrated that BAP domains with different primary amino acid structures and thus with different molecular weights can be 

used in the same experiment, providing the possibility of multiplexing. Alternatively to the changes in primary amino acid 

structure, the stable isotope format can also be used, providing another way to perform multiplexing experiments. Finally, we 

also demonstrated that our system could help to overcome another limitation of current methodologies to detect protein-protein 

proximity. For example, one can follow the state of a protein of interest at a defined time after its interaction with another protein 

has occurred. This application should be particularly useful for studying multistep intracellular processes, where the proximities 

between proteins and protein properties typically changed in a sequential manner.  

Conclusion: This approach has promised in adding temporal dimension in addition to helping reconstruct cell topology in space.  

Keywords: spaciotemporal structure, protein-protein proximity, spectometry 

 

 

 

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