別紙4 (課程博士)
Datc of submlsslon

平成Department

環1見29年

1月10日

SludentlD Numbel

生命工学専攻PP11Ca爪's name

氏名

学籍番号

第 093506 号

岡本吉晃

Supervlsors

手老青E吾

指導教員

岩佐精二

伊津野

真一

Abstract

論文内容の要旨(博士)Development ofne、¥ art所Cial biomembrane systems and theil' chal'acterization based on
Title ofThesis

mesoscopic m01'ph010gy and lateral di仟Usion

博士学位論文名(新規人工生体膜系の開発およびメソスケール領域における構造と側方拡散陛に基づいた特性評価)'琴︑玉(Approx.80o words)(要旨 1,200字程度)

ξ︑

Lipid bilayer is a fundamental structure of plasma membranes, and behaves as the reaction
field for various mem价'ane reactions through the 田'ganization of lipid domains and themolecule diffusion of biomolecules. The behavior of lipids and membrane proteins existing inIipid bilayerS 田'e important targets to elucidate the elementary processes of membranereactions. Therefore new experimental systems and methods for the 111embrane reactions aredemanded. supported lipid bilayer (SLB),、入lhich is one of artificia11ipid bilayer systems, hasbeen used as a ce11 membrane model to study the physicochemical properties of lipid bilay引',
and as a platform to investigate the structure and the function of membrane pl'oteins
Recently, graphene oxide (GO),、Nhich is one of the 即'aphene derivative with oxygen
functional groups such as hy山'oxyl, epoxy, and carboxyl groups on its surface, is exploitedfor the bi010gical application as a auorescence quencher 、¥ith unique properties. AISO
Synthetic auorinated lipids and surfactants which are substituted their hydrogen atoms in thehydrophobic alkyl chain X刃ith nuorine atomS 引'e applied in the field of biochemistry andmedicine for extraction and rem'ganization of the membrane pl'oteins according to bi010gicaland chemicalinertness of auorocarbons.1 expected that application of these materials, GOand auorinated lipids to dle s[B system leads to the develoP1τlent of ne、N methods andexperimental systems for the measurement of the behavior of biomolecules such as lipids andmembrane proteins in the lipid bilayer. Herein,1 Studied the development ofthe new artificial
Iipid bilayer systems using Go and a auorinated lipid for the measurement of biomoleculebehavior in Hpid bilayer.1 evaluate their struct1Ⅱ'es and physicochemical properties based on
the mesoscopic morph010gy and the lateral diffusion in sLBGraphene and Go have characteristic auorescence quenching abilities compared Xへlith oth引'general quencher molecules: independent of the wavelength of auorescence probes; 10ng
effective range. Go exhibits the amphiphilicity, because Go has a heterogeneously surface
Consisting of oxidized hydrophⅡic regions and nanoscale hydrophobic regions xNith grapheneStructure、 The fundamental study on the effects of GO X刃ith the microscopicaⅡy
het引'ogeneous surface on d〕e physicochemica1 御'operty of lipid bⅡayer is an imp0此ant
Subject for the application of Go in ule field of bi010gy and nanobiotechn010gy. As a first
Step of my work, we formed the s[B of l,2・dioleoyl・sn・glycero‑3・phosphocholine (DOPC)On Go by the vesicle fusion method. Go was prepared according to the modified Hummer'S
method, which is one of the common chemical exfoliation 1ηethods.1 evaluated the structure

and auidity of the sLB formed on Go by the atomic fm'ce 111i引'oscopy (AFM) and theauorescence recovery after photobleaching (FRAP) measurement.1 Proposed the structuralmodel ofthe sLB/Go system, and found that second layer of sLB 、Nhich is not formed by theVesicle fusion method generaⅡy,、Nas stably formed on GOTo evaluate the auidity of sLB formed on Go quantitatively,1 Conjugated quantuln dots(Qdots) as a bright auorescence probe to the sLB surface covalenuy f田' the single particle
U'acking (SPT) measul'ement.1 also introduced a lipid conjugated x¥ith polyethylene glyc01(PEG) chain on its head group in DOPC・S[B to suppress the non・specific adsorption of Qdotto sLB surface.1 0bserved the lateral diffusion of the Qdot・conjugated lipids in the sLB on
Go even under the effect of auorescence quenching by GO.1 found seV引'al Qdot‑conjugated
Iipids diffused between the Go region and si02 region, and evaluated the diffusion
Coefficient (D) of the Qdot conjugated・1ゆid in the Go and si02 re目ions on the basis of themean・square displacement (MSD) analysis. The average value of D of the Qdot、conjugated
Iipids on the Go region 、Nas appl'oximately 30少、o sma11er than that on the si02 region.1
mvestigated the morph010gy and auidity of sLB containing pEG・1ゆid on si02/si substrates
With and 、Nithout GO. Depression domains were observed in the sLB and incl'eased with theConcentration of pEG・1ipid on si02/si without GO, and the depression domains were
10calized on Go on the si02/si with GO. These results suggest that Go under s[BConcentrated pEG・1ゆid, and as a result D 、Nas sma11er in the Go region than in the si02
reglon/主巨The auorinated lipid affects to lipids and membrane proteins in a different manner n・om theCommon phospholipids with hydrocarbon chains.1 Prepal'ed the sLB of a novel partiaⅡyauorinated lipid l,2・di・(Π,11,12,12, B, B,14,14,14・nonaauorotetradecanoyb、sn、glycero‑3、Phosphocholine (F4・DMPC), and evaluated the latel'a11ipid mobility and its temperatul・edependence as the index to estimate the intermolecular int引'action of the partiaⅡy auorinatedIipid. Fluorination of lipid alkyl chain weakened the intermolecU1引・ interaction in thehydrophobic core in the lipid bilayer, and l evaluated its degl'ee qualitatively on the basis ofthe activation energy of lateral diffusion obtained 介0111 Arrhenius plots of D.
In these lNorks,1 Constructed new artificia11ipid bHayer syst引ηS applying Go andauorinated lipids, and revealed their structures and physicochemical properties.1 believe thatthese systems become a foundation for the experimental method for the highly accurate
measurement and for the contr01 0f the position, orientation, and activity of biomolecules in

Iipid bilayer me111branes.1 expect that the fundamental study of Hpid bilayer using the ne、Nmaterials such as Go and auorinated ⅡPid lead to greater understanding for the membrane

reacuon.1t is expected that the further development of these ne、N arti行Cia1 1ipid bilayer
Systems to reveal the elementary steps of the membrane reactions through the recognition of/'謡︑

the asymmetry of lipid bilayer, the detection of the microscopic change of the thickness ofIipid bilayel', and the expression of a ne、N function of membrane proteins