Advances in Planar Lipid Bilayers and Liposomes

322 Pages·2005·4.514 MB·1-323\322

by  A. Ottova-Leitmannova (Eds.)

CONTENTS
Contributors vii
Preface ix
1. Electrochemical Methodsand Their Application 1
Sławomir Kalinowski
2. Tethered BilayerMembrane Sensors with Small Transmembrane
Peptide IonChannels– Recent Developments, FutureResearch
and Potential Applications 49
Ping Yin
3. NMR Structure Determination ofProteinsin BilayerLipid
Membranes: The FXYD Family Proteins 77
Carla M. Franzinand Francesca M. Marassi
4. Cell SurfaceModelson PolymerSupports – From Artificial
Membranes toNative Cells 95
Motomu Tanaka
5. The Control ofMembrane Propertiesby Synthetic Polymers 121
Nickolay Melik-Nubarov and Oxana Krylova
6. Hydration Pressure and Phase Transitions ofPhospholipids 167
Helge Pfeiffer
7. ModelingProtein–Lipid Interactions: Recent Advances 187
Monique Laberge,Istva´n P. Suga´rand Judit Fidy
8. Modelingof BLMs inAspectsof Phylogenetic Development
ofVertebrates 237
Armen E.Zakharian and Naira M. Ayvazian
9. Electrochemical Study ofthe BilayerLipid Membrane 261
Erkang Wangand Xiaojun Han
v
vi Contents
10. Mechanismsof Membrane Permeabilization by Apoptosis-
Regulatory Proteinsof theBCL-2Family 305
OihanaTerrones, Aitor Etxebarria and Gorka Basan˜ez
SubjectIndex 317
CONTRIBUTORS
Naira M. Ayvazian 237
GorkaBasan˜ez 305
Aitor Etxebarria 305
JuditFidy 187
CarlaM. Franzin 77
Xiaojun Han 261
Sławomir Kalinowski 1
Oxana Krylova 121
Monique Laberge 187
Francesca M. Marassi 77
Nickolay Melik-Nubarov 121
HelgePfeiffer 167
Istva´n P. Suga´r 187
MotomuTanaka 95
Oihana Terrones 305
Erkang Wang 261
Ping Yin 49
Armen E. Zakharian 237
vii
PREFACE
The inspiration for lipid bilayer research, without question, comes from the
biological world. Although the first report on self-assembled bilayer lipid
membranes (BLMs) in vitro was reported in 1961, experimental scientists have
beendealingwiththeseinterfacialphenomenasinceRobertHooke’stime(1672).
Inthisconnection,theconclusionthatthefundamentalstructureofbiomembranes
isa lipidbilayerisbased on three pivotal experimentalfindings:
† Firstly, the elegant and simple experiment of establishing the orientation of
amphipathic moleculesatinterfacesby Langmuir in1917,
† Secondly, using Langmuir’s method Gorter and Grendel reported in 1925
that the extracted lipids from the plasma membrane of red blood cells (RBC)
thatoccupiedtheareaonthesurfaceofaLangmuirtroughwhichwastwicethat
oftheoriginalmembrane,andsuggestedthatitsstructureisnotunlikethatof
a soap bubble, and
† Thirdly, as deduced from the above indirect evidence, and from Fricke’s
electrical measurements of RBC suspension, as well as from the Unit
Membrane Hypothesis of J.D. Robertson based on electron microscopy in
the 1950s, the lipid bilayer structure was dramatically demonstrated after
D.O. Rudin and his associates reconstituted a black lipid membrane (BLM)
from the lipids extracted from cow’s brain (Nature, 1962, 194, 979).
Today, fromall lines ofresearch findings, all cell plasma membranespossess
a lipid bilayer structure, thereby underlying the lipid bilayer principle of
biomembranes. Thus, the lipid bilayer research over the past four decades has
evolved as an interdisciplinary effort, benefited by a cross-fertilization of ideas.
The lipid bilayer associated with life sciences and biotechnology is of current
interest to a diversity of investigators, including biochemists, biologists,
biophysicists, bioengineers and technologists, electrochemists, physiologists,
pharmacologists, surface and colloid scientists, and others working on ultrathin
films and membrane phenomena. In particular, BLMs have been used in a
number of applications ranging from basic membrane biophysics studies to the
conversion of solar energy via water photolysis, and to biosensor development
using supported bilayer lipid membranes (s-BLMs). s-BLMs (supported planar
lipidbilayers)providethefoundationforavarietyoflipidbilayer-basedmolecular
ix
x Preface
sensorsthataresensitive,versatile,inexpensive(i.e.,disposable)andopentoall
sorts of experimentation and development.
With the above background in mind, the present Advances series on BLMs
(planar lipid bilayers and liposomes) include invited chapters on a broad range
of topics, ranging from theoretical investigations, specific studies, experimental
methods, to practical applications. The author(s) of each chapter solicited to
focus mainly on the work of his/her laboratory, with minimal reviews of others.
Thegoalsofthechaptersarefornewcomers,butalsoforexperiencedscientists,
and for others who are not familiar with the research areas dealt with. An aim of
the series is to cover all aspects of lipid bilayer investigations, both fundamental
and applied. These contributed chapters are entities to themselves, thereby
strengthening the lipid bilayer principle of biomembranes, and are related to the
overall lipid bilayer venture. We are grateful to each of the contributors for their
expert knowledge and willingness to share generously their information.
H. TiTien
AngelicaOttova-Leitmannova
CHAPTER 1
Electrochemical Methods and Their
Application
Sławomir Kalinowski*
DepartmentofChemistry,UniversityofWarmiaandMazury,10-957Olsztyn,Poland
Contents
1. Introduction 2
2. Potentiometry 3
2.1. Measurementequipment 3
2.2. Originoftransmembranepotential 8
2.2.1. Donnanpotential 8
2.2.2. Diffusionpotential 11
2.3. Applicationofpotentiometry 13
2.3.1. Investigationsofelectricalprocessesincells 13
2.3.2. Lipidmembranesasionselectiveelectrodes 14
2.3.3. pH-sensitiveelectrodes 14
2.3.4. Potentiometricdetectionofmolecules 14
2.3.5. Investigationsofphoto-effects 15
3. Amperometryandvoltammetry 15
3.1. Potentiostats 15
3.1.1. 2-electrodepotentiostat 17
3.1.2. 3-electrodepotentiostat 19
3.1.3. 4-electrodepotentiostat 20
3.2. Applicationofamperometry 22
3.3. Applicationofvoltammetry 22
3.3.1. Measurementsofmembraneresistanceandcapacitance 23
3.3.2. Estimationofqualityofsolidsupportedmembranes 24
4. Chronopotentiometry 25
4.1. Galvanostats 25
4.2. Applicationofchronopotentiometry 25
5. Capacitancemeasurements 27
5.1. Characteristicsofthemeasurementmethods 27
5.1.1. Measurementswithasinusoidalsignal 28
5.1.2. Measurementsofcapacitancewithatriangularsignal 29
5.1.3. Bridgemethods 31
5.1.4. Pulsemethods 31
5.1.5. Compensationmethods 34
5.1.6. Conversionofcapacitancetofrequency 35
*Correspondingauthor.Tel.:C48-89-5233711;Fax:C48-89-5240408;
E-mail:[email protected]
ADVANCESINPLANARLIPIDBILAYERSANDLIPOSOMES,VOLUME2 q2005ElsevierInc.
ISSN1554-4516 DOI:10.1016/S1554-4516(05)02001-6 Allrightsreserved
2 S.Kalinowski
5.1.7. Measurementoftheminimumcapacitancepotential 37
5.2. Applicationsofmembranecapacitancemeasurements 37
5.2.1. Measurementofmembranethickness 39
5.2.2. Analysisofstabilityofbilayerlipidmembranes 39
5.2.3. Processofmembraneformation 39
5.2.4. Toxicologicalstudies 40
5.2.5. Investigationofphoto-effects 40
6. Referenceandauxiliaryelectrodes 40
6.1. Referenceelectrodes 40
6.1.1. Silver/silverchlorideelectrode 41
6.1.2. Calomelelectrode 42
6.2. Auxiliaryelectrodes 44
7. Protectionagainstnoises 44
References 45
Abstract
The chapter presents basic principles of the most common electrochemical techniques
usedforinvestigationsofbilayerlipidmembranes,andexamplesofelectronicequipment:
potentialmeters,2-,3-,and 4-electrodepotentiostatsandgalvanostats.Italsodescribes
the originoftransmembranepotentialin membranesystems,examplesforapplicationof
potentiometry, amperometry, voltammetry, and chronopotentiometry. An important and
characteristic parameter of membranes is their capacitance. Methods of capacitance
measurementandtheirapplicationarepresented.Duetotheirparameters,lipidmembrane
systemsrequireveryhighresistance,extremelylowcurrentsandhighcapacitance.They
are sensitive to distortions and external noises. Noise-protection methods are also
describedinthechapter.
1. INTRODUCTION
Life processes are dependent on electric phenomena taking place in cell
membranes. Transport of ions through the membrane and generation of the
membrane potential are among basic life processes. The development of
electrochemicalresearchmethodsallowstounderstandmanyimportantprocesses
observedincells.Thesemethods,knownfromclassicalelectrochemistry,turnedout
tobeveryusefulforanalysisofcellmembranesandartificiallipidmembranes.The
understandingofcertainmembranephenomenaenabledtheirpracticalapplication,
mainlytoanalyticalchemistry,medicine,andbiochemistry.Bilayerlipidmembranes
were found to be a good matrix for receptor molecules separated from natural
membranesandawidevarietyofartificialtransducers.
Electrochemical methods of analysis are relatively inexpensive and simple.
Measurement devices can be miniaturized and applied as portable instruments.
Rapid development of electronics makes it possible to employ numerous
electrochemical measurement techniques. The application of computers for
controlling measurement devices and data acquisition allows to use more and
ElectrochemicalMethodsandTheirApplication 3
moreadvancedelectrochemicaltechniqueswithsimpleelectronicanalogcircuits,
e.g., amperometry with programmable potential, electrochemical quartz crystal
microbalance (EQCM), impedance analysis. Many companies, e.g., National
Instruments, Advantech, CyberResearch, offer interfaces useful for data
acquisition and controlling external devices as relays, solenoid valves, electrical
motors, measurement offrequency,oscilloscope cards.
Measurement devices applied to membrane electrochemistry must be
characterized by certain important parameters like high current sensitivity in a
range of picoampers or lower, necessary for analysis of, e.g., single-channel
proteins. Very high resistance of membranes disturbs the measurement of the
transmembrane potential. Capacitance measurements performed using typical
capacitancemetersareimpossibleduetoatoohighmeasurementsignalapplied
to the membrane by the meter, usually higher than 1V. Bilayer lipid membranes
are broken by a much lower potential, oftenbelow200mV.
This chapter describes the equipment useful for membrane studies and
examples ofits applications.
2. POTENTIOMETRY
2.1. Measurement equipment
Measurementofthetransmembranepotentialisoneofthemostcommonlyused
andsimplesttechniquesinmembraneelectrochemistry.Figure1showsexamples
of the application of this kind of measurements to membrane systems. This
methodisgenerallyrestrictedtomembranesseparatingtwoelectrolytesolutions,
where the Donnan or diffusion potential is generated. Systems with membranes
covering solid support, e.g., metal, glassy carbon, conductive glass, do not
generatethis potential.
(a) (b) (c)
V V V
RE1 RE2 RE1 RE2 RE1 RE2
M C
M
Fig. 1. Examples of potentiometric measurements in membrane systems: (a)
membraneisseparatingtwowatersolutions,(b)themembraneissupportedona
gel electrode, (c) measurement of the membrane potential in a live cell using
application of a microelectrode. RE1, RE2 are reference electrodes, M is
membrane, C iscell.
4 S.Kalinowski
Correct measurement of the transmembrane potential in lipid membranes
depends on several conditions. Membranes sometimes exhibit resistance in a
range of a few dozen gigaohms. In this case the inputs of the devices used for
transmembranepotentialmeasurementsmustshowveryhighresistance,atleast
two ranges higher than the resistance of membranes. Another important
parameter is input offset current, whose value should be as low as possible,
lowerthan1pA.Otherwisethereadvalueofthepotentialwouldbeburdenedwith
agrosserror.Figure2presentstheimportantparametersofthesystemmeasured
andmeasurementdevicewhichhaveaninfluenceonthemeasurementaccuracy.
Sincethemeasurementwasperformedunderstaticconditions,thecapacitances
ofthismodelcircuitcouldbeignored.Themembrane,electrolytesandelectrodes
are shown as a source of the potential E composed of the membrane potential
M
(Donnan or diffusion potential) and electrode potentials on both sides of the
membrane. The membrane with electrodes and electrolytes exhibits the
resistance shown in Fig. 2 as R . The membrane resistance is usually several
M
ranges higher than the resistances of electrodes and electrolytes. In this case it
can be assumed that R isequal to themembrane resistance.
M
Measurementdevicesdonothaveidealinputparameters.Themainsourceof
measurementerrorscanbetheinputresistanceofthemeasurementdeviceR
inp
andtheinputoffsetcurrenti .TheinputresistanceR causesadecreaseinthe
off inp
readvalueofthepotential.Additionally,theinputofthemeasurementdeviceisa
sourceoflowdirectcurrent.Inthecaseofmeasurementofthepotentialgenerated
by a signal source with high internal resistance an additional value appears,
causing a measurement error. The input offset current i generates additional
off
voltage added to the measured signal.
If the input offset current i is ignored, the read value of the potential can be
off
described by the equation:
R
E ZE inp (1)
P MR CR
M inp
Signal source Measurement device
RM in
E Rinp EP
M
Fig.2. Essentialparametersofthepotentialsourceandthemeasurementdevice
affecting theaccuracy ofthemeasurement.E isthemembranepotential,R is
M M
themembraneresistance,R istheinputresistanceofthemeasurementdevice,
inp
i is theinputoffset current, E isthe displayed value ofpotential.
off P
ElectrochemicalMethodsandTheirApplication 5
IfthemembraneresistanceR isequalto1010U,theinputresistanceR equal
M inp
to 1012U results in a measurement error of about 1%. In practice, the input
resistanceofthemeasurementdeviceR shouldbeintherangeof1013–1014U.
inp
Theinputoffsetcurrenti causesanadditionalerrorDE whichcanbecalculated
off off
from Ohm’s law:
DE Zi R (2)
off off M
ForthemembraneresistanceR equalto1010Uandthecurrenti equalto1pA
M off
the error DE is equal to10mV.
off
Manydevicesdesignedforvoltagemeasurementareavailableonthemarket.
Their suitability for membrane applications depends on the input resistance R
inp
andtheinputoffsetcurrenti .Simplevoltagemetersareuselessduetotheirlow
off
input resistance, usually ca 1MU. The best devices for membrane potential
measurement are electrometers – instruments with extremely high input
resistance,reaching1015U,andinputoffsetcurrentintherangeoffemtoampers.
AlsopH-metersandion-meterscanbeusedforthemeasurementofpotential,as
theyhaverelativelygoodparameters.pH-metersaredesignedforco-workingwith
glass electrodeswith internal resistance ina range ofhundreds ofmegaohms.
The input resistance of the device is usually given in the technical
documentation attached to the device. The input parameters can be estimated
bymakingseveralsimpleobservationsandmeasurements.Ifthedevicedisplays
stability, the voltage equal to zero or close to zero usually indicates low input
resistanceofthedevice.Deviceswithhighinputresistancearesensitivetostatic
electrical fields occurring, e.g., when clothes pick up static electricity. Keeping a
hand close to the wire connected to the input causes changes in the displayed
voltage.Itisimportanttobecarefulbecausethe inputscanbe destroyedby this
electrical charge.
The input resistance can be measured using a 1.5V battery and a reference
resistor of the same range as the input resistance. First it is necessary to
measure the voltage E of the battery connected directly to the input (Fig. 3a).
NexttheresistorRshouldbeconnectedinserieswiththebattery(Fig.3b)andthe
voltageE0shouldbemeasuredagain.TheinputresistanceR canbecalculated
inp
(a) Measurement device (b) Measurement device
R
E 1.520 E 1.155
Fig.3. Estimationoftheinputresistanceofapotentialmeter:(a)batteryvoltage
measurement,(b) voltage measurement with resistor Rconnected inseries.