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Mar 1, 2010 (Vol. 30, No. 5)

Protein-Protein Interactions

In-Depth Investigation of These Complex Relationships Is Increasing Knowledge Base

  • Biophysical Methods

    Click Image To Enlarge +
    EIN (blue) and HPr (light brown) form a stereospecific complex with an affinity of ~5 µM. (University of Missouri)

    Protein-protein interactions are really driven by biophysical forces, namely electrostatic forces, van der Waals forces, and polar bonding between amino acids at the interface between the interacting proteins. Biophysical methods of all kinds are popping up everywhere to study these forces and investigate protein-protein interactions.

    Chun Tang, Ph.D., assistant professor at University of Missouri, studies protein dynamics using nuclear magnetic resonance (NMR). He has developed a new NMR method for this purpose that is based on 2006 data published by his lab that shows that, when two proteins interact with each other, they form not only a stable complex, but also a myriad of other conformations or intermediate states.

    “We use a technology called parametric-relaxation enhancement to detect those intermediate states,” explained Dr. Tang, who added that a bacterial phosphoryl transferase protein complex called EIN was used as the model for the study. The structures of these proteins in EIN are already known. Based on these structures, Dr. Tang was able to build a picture of the dynamics of the complex.

    “With this new technology, not only can we see the interaction between components in the complex, but we can also get a timescale for its formation. In the NMR field, we not only want to know protein structure, but also the protein dynamics, to see how the proteins fluctuate between functional and intermediate states. This new technology will provide information on that fluctuation.”

    In the same vein of biophysical approaches, Wyatt Technologies teamed up with Genentech to conduct comparative studies of composition gradient static light-scattering (CG-MALS) as a complementary and orthogonal technique to surface plasmon resonance (SPR). The two technologies were compared as they were used to analyze interactions between an antibody-based biopharmaceuticals and both its target receptor and an effector protein.

    “Biopharmaceuticals are complicated molecules,” noted Daniel Some, Ph.D., principal scientist at Wyatt Technologies. “For example, they may be multivalent. SPR does not give you the whole picture because it does not confirm multivalency very robustly, but by adding in the CG-MALS measurement we can clear up some of the additional questions and provide a more complete picture.”

    According to Dr. Some, composition gradient static light scattering is a technique that does not require immobilization but is label-free and in solution. “Often, things happen in solution that do not occur when one of the components [of an associating protein complex] is immobilized onto a surface, as is required by SPR.”

    Due to the fact that CG-MALS does not require immobilization of protein components, it has become a powerful technique for distinguishing between solution-based effects and immobilization-based effects in the study of protein-protein interactions.

    Protein-protein interactions were, for most of the lifetime of molecular biology, an enigma. However, with more powerful genetic, bioinformatic, biophysical, and computational methods, solving the “molecular” puzzle has become a little easier.

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