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Creation of Membrane Stress Biotechnology

In biofunctional materials design laboratory, we are pioneering a new academic field named as “Membrane Stress Biotechnology (MSB)”. A biomembrane is a frontier of the biological cell against its environment and a functional interface to make cellular responses for its survival. When the biological cells were exposed to the environmental stresses, such as heat, pH, and oxidative stress, the cell responds to them or falls into death. It has gradually been revealed that the biomembrane itself can make various responses against such stresses (i) by recruiting the environmental elements on its surface and (ii) by creating various kinds of supramolecular structures with highly-ordered state. A key to establish the MSB is to understand the principle of the potential functions of biomembrane induced by stress application. One can apply it for the design and development of the functional material/ process based on the “Break-down” or “Build-up” approach and one could clarify the molecular mechanism of the potential functions of biomembrane based on the “Build-up” approach, resulting in the establishment of the method of the diagnosis and remedy of Alzheimer’s disease or hemodialysis Amyloidosis. The conventional knowledge, such as Genome and Proteome, is not clear enough to achieve the above purpose. It is really important and needed to systematize the information of the membrane and its stress-response dynamics. “Liposome”, a closed phospholipid bilayer membrane, can be regarded as a LEM-Unit (as shown in Fig.) which plays a main role as a core material in “Membranomics” research.

Attractive Potential Function of Liposome Revealed under Stress Condition

The liposome has conventionally been utilized as a model biomembrane. It has been reported that the liposome could induce a variety of “potential” functions under the stress condition. A molecular chaperone-like function is one of the potential functions; the liposome could interact with the protein at abnormal conformation and could refold and activate it just like as a natural molecular chaperone (GroEL/GroES). The key parameter of the potential function has been shown to be strongly dependent on the “local hydrophobicity/hydrogen bond stability” of both protein and liposome under stress condition through the analysis using aqueous two-phase partitioning method, immobilized liposome chromatography, and dielectric dispersion analysis etc, showing that the simple liposome surface could recognize the conformational state of protein and interact with it. It has been shown that such basic findings can also be applied to the other kinds of potential functions such as protein translocation across the membrane and membrane fusion, depending on the intensity of the fluctuation of protein and membrane under the stress condition. The above-described functions of liposome are expected to be utilized for several diseases caused by “Membrane-Abnormality”, such as Alzheimer’s disease.

LIPOzyme Chemistry：Material Design Based on Liposome

Fundamental Technology in Biomembrane Process Chemistry

LIPOzyme (Liposome+Enzyme) function can be defined as the enzymatic function of liposome itself induced under the variation of the environmental stress; (i) molecular recognition function, (ii) function to convert the protein conformation, (iii) catalysis for bioconversion and so on. Based on the LIPOzyme functions, several kinds of catalysis have been designed such as (i) cholesterol oxidase-like function of Ab/Cu complex, (ii)antioxidative SOD/CAT catalysis in one-pot, and (iii) hydrolytic enzyme-like catalysis (like as a-chymotrypsin). Surprisingly, the liposome could recognize and recruit the minimal elemental materials (such as peptide fragments, ligands, and so on), which have negligible activity, and could furthermore induce the enzyme-like function (LIPOzyme function), which is equivalent to the natural enzyme, by the reconstruction of the supramolecular structure of the elements. It is expecting that the LIPOzyme functions of liposome could be extended for the creation of novel “biomembrane process”, which can recognize(separate), catalyze(convert), and transport the substrate materials and their products through the rational modulation of various kinds of dynamic interactions on its surface, such as hydrophobic interaction and hydrogen bond stability, similarly in the case of the natural biomembrane. The above findings show that the bio-inspired materials could be prepared by using the liposome as a core unit

Prospectus of Membrane Stress Biotechnology – Challenge to Alzheimer’s Disease

In order to establish the MSB, a“Life-Environment Minimum Unit (LEM-Unit)” should herewith be defined as an indispensable and minimal unit of “life” and a complex coupling “life” and “its environment”. The minimal requirement of the LEM-Unit is for it to harmonize with other LEM-Units in the global environment and to integrate them as its sub-unit. Although there are many kinds of components of biological cell as a fundamental unit of “life”, among of them the “liposome (model biomembrane)” itself, a closed bilayer membrane, is indispensable one regarded as the “LEM-Unit”. This is because the liposome could intake “water” as an indispensable environment on its surface and in its inner space. We therefore employed the “liposome” as “LEM-Unit”. It has gradually been revealed that the essential characteristics of biofunctions are highly dependent on the functionality of the active interface (biomembrane), which can transform itself dynamically, based on the experimental findings on LIPOzyme. LEM-Unit can interact with other units because of its intrinsic“fluctuation nature” which makes it possible to respond against the subtle variation of environmental energy. We can re-define the liposome as “Self-Evolving” or “Self-Driven” Unit harboring the functions to convert the material, energy, and information. Such a liposome could induce the formation of various kinds of ordered structure or pattern through the non-linear/non-　equibrillium dynamics.

It has been believed that the conformational abnormality of the specific peptide could be a key subject in Alzheimer disease and Hemodialysis Amyloidosis. On the contrary, we consider that the abnormality of the biomembrane itself could be a key of the conformational abnormality of Ab and are also challenging to the design and development of the diagnosis and cure system of the “membrane abnormality” by using the “LEM-Unit”.

References (main papers in recent 3 years)

(1) Hiroshi Umakoshi, Keishi Suga, Huong Thi Bui, Masato Nishida, Toshinori Shimanouchi, and Ryoichi Kuboi, Charged Liposome Affects the Translation and Folding Steps of in vitro Expression of Green Fluorescent Protein, J.Biosci.Bioeng, 108(5), 450-454 (2009)

(2) Toshinori Shimanouchi, Hiroshi Umakoshi, Ryoichi Kuboi, Kinetic Study on Giant Vesicle Formation with Electroformation, Langmuir, 25(9), 4835-4840 (2009)

(3) T.Shimanouchi, E.Oyama, H.Ishii, H.Umakoshi, R.Kuboi, Membranomics Research on Interactions between Liposome Membranes with Membrane Chip Analysis, Membrane, 34 (6), in press (2009)

(4) T.Shimanouchi, P. Walde, J.Gardiner, S.Capone, D.Seebach, R.Kuboi, Inversion of the Configuration of A Single Stereocenter in A b-Heptapeptide Leads to Drastic Changes in Its Interaction with Phospholipid Bilayers, ChemBioChem, 10, 1978-1981 (2009)

(5) Huong Thi Bui, Hiroshi Umakoshi, Kien Xuan Ngo, Masato Nishida, Toshinori Shimanouchi, and Ryoichi Kuboi, Liposome Membrane Itself can Regulate Gene Expression in Cell Free Translation System, Langmuir, 24(19), 10537-10542 (2008)

(6) Le Quoc Tuan, Hiroshi Umakoshi, Toshinori Shimanouchi, Ryoichi Kuboi, Liposome-Recruited Activity of Oxidized and Fragmented Superoxide Dismutase, Langmuir, 24, 350-354 (2008)

(7) Hiroshi Umakoshi, Kengo Morimoto, Yuji Ohama, Hideto Nagami, Toshinori Shimanouchi, Ryoichi Kuboi, Liposome Modified with Mn-Porphyrin Complex Can Simultaneously Induce Antioxidative Enzyme-Like Activity of Both Superoxide Dismutase and Peroxidase, Langmuir, 24, 4451-4455 (2008)

(8) Makoto Yoshimoto, Yuya Miyazaki, Ayumi Umemoto, Peter Walde, Ryoichi Kuboi, Katsumi Nakao, Phosphatidylcholine Vesicle-Mediated Decomposition of Hydrogen Peroxide, Langmuir, 23, 9416-9422 (2007)

(9) Seiichi Morita, Yuya Hamano, Ryoichi Kuboi, Effects of Fatty Acids on Interaction between Liposome and Amyloid b–Peptide. Maku (Membrane), 32, 215-220 (2007).

(10) Toshinori Shimanouchi, Peter Walde, James Gardiner, Yogesh R. Mahajan, Deter Seebach, Anita Thomae, Stephanie D. Krämer, Matthias Voser, Ryoichi Kuboi, Permeation of a b-Heptapeptide Derivative across Phospholipid Bilayers, Biochim.Biophys.Acta, 1768, 2726-2736 (2007)