RIKEN

Conformation analysis and NMR screening with NMR facilities
of the world’s largest scale

By utilizing the structural information in the process of developing new drugs, it is expected to shorten the development period, decrease the cost and the side effects. Particularly, the interaction analysis between targeted proteins and compounds using NMR is a strong analysis tool in the drug discovery research.

In NMR platform we retain about 40 high-throughput NMR apparatuses (900MHz: 3; 800MHz: 14; 700MHz: 6; 600MHz: 17) to analyze protein conformations and their function, and have technologies that can determine more than 300 protein conformations annually. We systematize the interaction analysis between targeted proteins and compounds based on the technologies that can observe signals of proteins (molecular weight 40,000-50,000) and those of compounds (molecular weight about 100,000). NMR interaction measurement techniques and sample preparation techniques are developed to systematically promote the process of interaction analysis. Moreover, we combine it with NMR conformation analysis pipeline to construct a support system for a series of screening with NMR from sample preparation of targeted proteins to protein conformation analysis and interaction analysis of compounds in order to support individuals involved in drug discovery within and outside Riken.

To promote drug discovery research, Riken develops two technology platforms of conformation analysis and NMR screening to support various drug discovery researches through exploration of targeted molecules and support of screening.

Lead compound screening platform using interaction analysis with NMR

Structure-Based Drug Discovery (SBDD) for drug design and optimization based on targeted protein conformation is a representative methodology in the drug development process utilizing conformation information. Particularly, a procedure that constructs molecules from small molecular building blocks has been focused on recently. As the compounds found by this methodology are assumed to have weak interaction with targeted proteins of a new drug, with the dissociation constant ranging from mM to mM, especially in the initial phase, combination it with NMR possessing strengths in interaction analysis within this range is considered to be markedly effective. In the NMR platform, we develop two techniques of “NMR interaction measurement technique” and “sample preparation technique” as well as two systems of “standardization and automation of working process” required for systematic analysis of interaction between targeted proteins and compounds in order to establish elemental technologies for interaction analysis with NMR. We also construct a lead compound screening platform utilizing interaction analysis by combining it with the elemental technologies

In the development of NMR interaction measurement techniques, development and sophistication of sensitive NMR interaction analysis techniques are pursued to identify the binding site and to optimize the structure based on chemical shift perturbation and crossrelaxation, utilizing characteristics of NMR with strengths in analysis of weak interaction ranging from mM to mM (Fig.1). In the development of sample preparation techniques, techniques to analyze interaction under various conditions of samples are produced by developing three types of techniques to prepare analysis samples such as: (1) high concentration of low-soluble potential compounds; (2) technology to introduce stable isotopes having complex and selective patterns into targeted proteins as an advanced labeling technology; and (3) a sample preparation technique for analysis utilizing the residual dipole-dipole interaction (RDC) and the paramagnetic relaxation enhancement (PRE) to use information on the distal distance. Furthermore, we review the optimal data format to describe the research process, and develop a data management system to achieve information management on the history of samples and data, sample tracking, and data mining. Moreover, working process is standardized and automated to establish elemental technologies by improved reproducibility and constructing an automated apparatus for concurrent multiple sample analysis.

Furthermore, we construct a lead compound screening platform (Fig.2) that screens compounds with strong interaction by exploring the weak interaction between proteins and low molecular compounds and binding several molecules with weak interaction together, using interaction analysis pipeline with NMR that combine elemental technologies

Construction of unique NMR platform and promotion of external use

Utilizing the results of previous researches, NMR platform has constructed “NMR conformation analysis pipeline” (Fig.3 )that assesses the compatibility in NMR analysis of protein samples, prepare stable isotope labeled protein samples, determine multidimensional NMR data, and decide the protein conformation based on measured data. It has been already available to those outside Riken. By combining this pipeline with NMR screening platform a unique and powerful NMR platform can be constructed. Riken Yokohama Institute provides the external use of NMR conformation analysis pipeline in three forms: use as a task for “Open Advanced Facilities Initiative for Innovation” (non-proprietary, supported by the government), proprietary use (fee-based), and non-proprietary use (fee-based with a discount).

About NMR Facility, Yokohama Institute, RIKEN, click here.