3TP3-06 Streptavidin 2D crystals as solid supports for the visualization of biomolecular processes by high-speed AFM(The 47th Annual Meeting of the Biophysical Society of Japan)
3TP3-06 ストレプトアビジン二次元結晶基板の高速AFM観察への適用(バイオイメージング,第47回日本生物物理学会年会)

Daisuke Yamamoto, Naoki Nagura, Saeko Omote, Masaaki Taniguchi, Toshio Ando
2009 Seibutsu Butsuri  
TheBiophysicalSociety of Japan General IncorporatedAssociation Gene expTcsgion ig probably ]ntegrated into architectural "amcwork of Lell nucleus How is dynamiL change in the mtranuclear stmctuTeg corrL]atLd with nuclear funcuoning i Presently it is diliiLult t{} answer the quesuon well because intnnuclear structure is not wLll ebserved with consentional mieiogeopy WL therefore dcvdoped pupil proJeLtJon apodized phage contragt (PPAPC) miuosLopy PPAPC is a combination oi apodiiLd phase Lontratst
more » ... (APC) and pupil piqect]on phase Lontrdst (PPPC) Using PPAPC we observed fme structures within the interphase nuLleus in 1rvmg Hela cells without staining The fJne structures "ere mo-ng dynamically withm the nucleus Simultaneoug observation of the nucleus "ith PPAPC and xvth fiuorescence sho"ecl that parucles s-ere reall) moimg mside the nucleus Te identifv the moNmg paiticles in the nucleus -L intieduLed mteractue obscrvaLion of optLaT and eleLtron rnicroscopy We reeoided particle movements in thc nudeug vv ith PPAPC miLrosLepe v" tth optical slice thiLknLsg of 700 gOOnm Dunng the obsenat]on we appliLd fixative to the cell under the opL]cal micioscope The cell "as fixed within 100msLL attu the apphLatien of fixatave ThL idLntieal nuLTeub was sliLed sequentaally vLith thickncss of 70nm and obscrvcd vLith elcction miLrosL{)p> 10 12 sequential imagLs oi eleLtron micrographs revealed the movmg partiLILs m the tLvmg nucleus obser}ed with PPAPC Mechanism of the inttanuclear movement "i]] also diiLusbed in this pregentatlon 3TP306 XFVVF7t--=ptitva,EasfiopK-gAFManaj"opfiM Streptavidin 2D ciygLals As sohd supports for the visuahzation of biomolecular processes by high spLed AFM Daisukc YAmamoto (1) Ndeki Ndsur L{2) 5dcko Omote (2} Masaaki LamguLhi (2) Toslvo Ando (1) (Cl) Coilege qf Sctence and kngtneLnng Kanacawa Umversiij CR[5evJSf f2) CotltsL t)fSfTLn(e and FngTneering Kanazawa Vniveistty} For the direct observation of biomoleLular proeLssLs by atomic force microscopy (AI7M) sohd bupports are required to fulfi11 sevLial Lntuia buLh tis surface IlatnLgg a eapabihty to LontroJ the orientataon of the immobihiLd moleLules and abscncc of nongpeciiie protein adgorption Here sve charactenzed gtTeptavidin two dimenbional Lrystals as substrate surface In this study tlllee types of streptavdin crystals "ith C222 P2 and Pl symmetnes were obtained on thc supperted 1ipid bilayeis The surface ot these gLrepLavidin Lrystals ib flat enough to idLntity the indiv]dual meleeules on these suppoits ProtLms were speciftcally immobilized through streptavidin biotm 1]nkage on the crystal suitaceg whilL nenspeufiL adsorption of non biotm}lalLd protuns wls highl) restncted as rLvLalLd b) flueresLent miciobLopy These propertieg of thc strLptavidiii erystal surfaces ensurL thL visuahiabon ef spe"fJLally nnmobdized proteing in a muJt] component sample ImpoiLintly thL inLrt burfwes towards nonspecitic protein adsorpuon pegsibly guarantLL LhL immebilized protems against disfunLtion As test samples calumodulin (CaM) GroEL GroFS and actm filamentg "LrL ebser}ed by high speed AFM The contormational LhAnge ef CaM upon bmdrng lo calcium wuag dLteLtLd ds d Lhange in height of this protLm Furthermore Lontrolled o"entation of immobihLed GroEL GroE5 Lomplex was achieved by conlug lting biotm dt aspeclfic slte of (JroEL Finallv 1poi)meri7ation process of a smgte actm filament "as traced 3TP401 7tF>i7t-xf:S67kxxdipta)7Sleeo"+reee Opucal Trappmg of Arninu Aadvn Aqueous Solutron by Photon Force Tdtsuya ShoJL(1) Yasuyukd TKuho (1) Nebeiu Kitamuia ") ((1) llokhatdo Untv ) The opticab1 trappmg teLhmque would bL thL only methodology in a non contact mode to Lapture and 3D mampulatL a molLcu]L in solution phase Manipulalton ot bioaLkvc tunLtienal moleLules hds potential applicabons m bietmedical science The target of rnolecular trappmg hag hitherto been 1imited only to a few large rnacromolccular gystcms Naturd] polymers such as DNA and artifiual polymers (Y Tsuboi M Nishmo Y Matgue K ijiro N KitarnuTa Bull Chem So( IPn Vol 8e (2007) 1926 1931) SmaltmoleculcshavLneverbeenopticallytrnpped m sotution but instead thLir tranglat]onal diffubion has beLn obgerved to be gupprLssLd at atoLcd point due to a shallow optical potLntial In the present study we exdmined optical trapping ot sLveral ammo auds in aqueous CD20) solution by focusing a near infraiLd CW ]dser beam Photon torce induced moleLular issemblLng of amino acids was rLvet]ed by means of mi"oscopiL imagmg and Raman m]LrospeLtrescepy We expemncntally demonstrated that photon forLe Lan trap dmmo acids d]sgolved in aqueous solutaons Amino aL]d moleculLs assLmble in a fevv mmutes at lhe tocnsed drea A gro"th rate ot aggregate mcreages with mereases in Loncentrataon ind laser power The ongin of the present optical trappmg ot ammo auds wab ascribed to the c]ugtLT trdppmg mcchan]sm (Y Tsuboi T Shnyi N Kitamura Jlpn j Appt Ph)s Vol 46 (2007) ] l234 L1236) and thL tLchnigue Lould be extended to other smdlt molecules or srnd]] oligmers Such molecular trappmg and manipulauon based on photon force has a "ealth oi apphcation buLh as selective drug dehvery to a 1iimg cell moleLuTar separatton targeted synthLsis ef bioaLtrve molecules (protcin) Formation ol an epithehal tube of ging]L eel] tsize circumference
doi:10.2142/biophys.49.s62_1 fatcat:yswdkl5wgjc6jml3e274zo27oi