Live cell, a fundamental of all living systems, consists of many different types of molecules whose size, structure, property, and function are varied. Dr. Itaru Hamachi has been interested in protein molecules in particular, and challenged to develop new chemistry that allows selective labeling/modification of endogenously expressed (natural) proteins under multimolecular crowding conditions such as live cells. He originally invented "Ligand-directed chemistry" for selective labeling of endogenous protein. This is a new chemical strategy based on coupling molecular recognition with proximity-driven chemical reaction, which allows for selective modification of natural proteins without genetic engineering in live cells, tissues and in vivo. He also discovered a supramolecular approach for selective protein imaging on the surface or inside of live cells. Furthermore, he has showed strong interest in creation of novel semi-wet supramolecular materials, by mimicking cellular structures and functions. He has made pioneering contribution to create novel functional supramolecular hydrogel materials comprising multiple components with dynamic ordering and response.
1 Live cell organic chemistry for proteins
Traditionally, proteins analysis has been conducted using purified samples in dilute aqueous solution in most cases. However, it is now being recognized that the structure and function of natural proteins in live systems may be rather different from those under such pure conditions. In order to analyze and control protein structure and functions in natural habitats, Hamachi developed a new method which enables to chemically label natural proteins in live cells and in vivo. This method relies on the proximity-effect, where the selective molecular recognition can greatly accelerate the subsequent chemical reaction with a target protein, termed ligand-directed chemistry (LDchem). In LDchem, a reactive and cleavable linker is designed that connects a ligand for selective recognition to a protein-of-interest (POI) with a probe to be tethered to POI. To date, his group discovered several efficient chemistry using tosyl, alkoxyacylimidazole, dibromophenylbenzoate, as the reactive linker. He has demonstrated that LDchem allows for the protein-specific labeling and imaging with high selectivity in live cells, as well as cell lysates and in pure test tubes. The target proteins are being now extended by appropriate selection of the ligand and the reactive linker, from intracellular proteins such as carbonic anhydrase, FKBP12, and HSP90 to membrane-bound proteins such as folate receptor, G-protein coupled receptors (GPCR) and neurotransmitter receptors (Glutamate receptors and GABA receptors) on live cell surfaces. The chemically labeled GABAaR, for example, was successfully converted to a fluorescent biosensor in live cells, which was used to give new drug candidates for GABAaR from high throughput screening of small molecules library. He also developed a novel Zn(II)-dependent proteomics approach, by designing a series of protein labeling reagents that can be activated by Zn(II) ion. This is a novel concept, that is a conditional proteomics, for unveiling roles of biological metal ions in chemical biology. In addition, he developed a chemogenetic method based on molecular recognition using coordination chemistry, which is able to regulate a native function of AMPA receptor, one of the Glutamate receptors, in live cultured neuron, as well as in model cells.
2 Supramolecular approaches for live cell protein imaging
In 2009, Hamachi discovered a series of ligand-probe conjugates are self-assembled to form nano-aggregates which were collapsed upon binding to a POI. He clearly demonstrated that this recognition-driven disassembly took place not only in pure test tube conditions, but also in cell surface and even inside of live cells. This principle allowed development of new supramolecular strategy for live cell fluorescent and/or NMR imaging of a target proteins in a turn-ON mode with low background signals.
3 Cell-mimic semi-wet supramolecular materials consisting of multiple components
Live cell is not disordered crowding systems consisting of many different types of molecules, but rather spontaneously well-ordered soft materials that contain different types of orthogonally assembled molecules and distinct spaces. Hamachi sought to mimic several essential factors of live cells by synthetic supramolecular assembly and succeeded in creating semi-wet soft materials made of supramolecular hydrogels. He visualized the self-sorting events of two different supramolecular fibers in situ and real time manner for the first time, by confocal laser scanning microscopy.
Also, he constructed multiple components systems consisting of supramolecular hydrogels embedding mesoporous silica nanoparticles, enzymes and proteins, layered clay, a synthetic amplification cascade, and fluorescent probe molecules, some of which were demonstrated to function as novel logic-gate responsive sensors, biomarker-sensitive controlled release matrix, and rheologically fine-tuned soft-materials. Such multiple-component materials systems are highly pioneering, which can provide a novel guidance to design functional soft-materials.
Dr. Itaru Hamachi largely stepped out of the conventional organic chemistry framework using purified molecules in pure dilute solution systems of test tubes and pioneered a new field of organic chemistry and supramolecular chemistry in multimolecular crowding aqueous environments. His enormous contribution and achievement in this really interdisciplinary research field between chemistry and biology are undoubtedly worldwide and invaluable, therefore, he deserves The Chemical Society of Japan Award