Electrochemical behaviour of natural and biosynthetic polynucleotides at the mercury electrode. VI. Influence of the base composition on the electrochemical behaviour of natural and biosynthetic polynucleotides

Bioelecioclrentistry

and

Bioenetyetics

3, 462-473

(1976)

Electrochemical Behaviour of Natural and Biosynthetic Polynucleotides at the Mercury Electrode. VI. Influence of the Base Composition on the Electrochemical Behaviour of Natural and Biosynthetic Polynucleotides * by J. 31. SEQUARIS, Institute Centre

of Chemistr)-. Jiilich

(KFA),

B.

Institute Federal

P. VALENTX

MALFOY, 4, Applied Republic

Physical

and H. IV. NURXBERG

Chemistry,

Nuclear

Research

of Germany

The electrochemical behaviour of DNA of various origin and of biosynthetic polynucleotides at a charged mercuryjsolution interface was studied in relation to their respective composition in bases. Two independent methods, phase sensitive ax. polarography and single sweep voltammetry, at a hanging mercury drop electrode were used to ensure the reliability of the results_ As all the biopolymers studied had a doublestranded conformation, the unwinding of these double-strand form due to the adsorption forces and to the electric field in the interface is indicated. The height of the reduction peak in single sweep voltammetry and its time integral vaq- linearly with the percentage of the bases A-T (adenine-thymine) in the biopolymer. The variation of the height and of the potential of a further capacitive n-c. peak shows that the phenomenon of reorganisation at a charged surface is also correlated to the nature of the bases. These results emphasize the importance of the base composition of the biopolymer as well as of its molecular weight and its conformation (single-stranded or double-stranded) in the course of niteraction with a charged surface.

Introduction We have shown 1~”previously by sweep voltammetry at the H.M.D.E. that, after adsorption, native calf thymus DNA is reducible in a totally irrel-ersible electrode process from an acid solution (pH 4 to 7) to very strongly adsorbed products forming a blocking film on the electrode l

Presented

chemistry.

Jiilich,

in part 27-31

at

the

October

3rd 1975.

International

Symposium

on Bioelectro-

462

Sequari,

Malfoy,

Valenta

and Niirnberg

surface and acting as catalysts for hydrogen evolution. The reduction occurs in the same potential range as for adsorbed denatured D3l-43~~ and is due to the electron and proton uptake by the accessible adenine and cytosine moieties. accordin g to a mechanism established earlier b> JXSIK and ELVISG” for the isolated bases adenine and crtosine and the correspoCding mononucleotides. Our findings on the adsorption and reduction are further confimled by x-eqr recent n.c. polarographic measurements at the D.3I.E. of MALFO\- and REYX-ACD.~ Further we have shownlz’ that a prerequisite for the reduction of adsorbed native DNA is the

unwinding of its double helical regions similar to that in which denatured DNA

into the single stranded is immediately adsorbed.3

stage This

interfacial unwinding occurs under the action of the adsorption forces and is also significantl>- fal-oured b>- the interfacial electric field at the charged interface. Meanwhile our findings in this respect have been further confirmed br the results obtained by PALECEK~ with nomlal pulse polarograph)- at the D.M.E. In a preceding papers we reported on our detailed and estended studies on the sequence of interfacial events during the adsorption time at \-arious adsorption potentials and developed a model of the comples interfacial behaviour of native DNA consistent with all collected esperimental evidence_

It is well known that the stability of helical regions largely depends on the composition of the base pairs linking both complementary strands aict hydrogen brigdes, according to the WATSOS-CRICK rules. Regions rich in adenine-thymine pairs (X-T) are more labile than those where guanine-cytosine pairs (G-C) prevail. As LEWIS~ has shown this is connected with lower dehydration eneragain in X-T-pairing compared with the formation of G-C-pairs. Thus, it is to be expected that native DNA and related biosynthetic polynucleotides o f different origin w-ill show quantitati\-e differences in electrochemical behaviour due to varying composition with respect to the percentage of paired bases of different nature, although the global qualitative behaviour will be similar to that of calf th>-mus DNX. The present paper is concerned with the investigation of those effects and also with the influence of varying molecular weights. These problems have been hitherto only tcuched in a few sporadic in\-estigations on denatured DNA’0 and on the kinetics of enzymatic degradation of native DNA from \-arious origins, 11,1” but were not \-et approached in a svstcmatic manner_

Experimental

The DNA-types and the bias)-nthetic polynucleotides studied are listed with their main physicochemical characteristics in Table I. The substances were either purchased or prepared according to I<_~Y et nl.13 for calf thymus DNA and to M_ARMUR~~ for bacterial DNA. All DNXQ-pes should ha\-e qualitatil-el!the same polarographic behal-iour. The)-

Electrochemical

Behaviour

of Natural and Biosynthetic ‘Polynucleotides

$63

were also not influenced by treatment with phenol to remove the last Other chemicals were of reagent or even MERCK traces of protein. “Suprapur” grade.

ami?Mefl'Iods

Appnratzcs

I. General. - Measurements of sedimentation coefficients were carried out with an analytical ultracentrifuge BECKMAX E. The molecular weight was evaluated with the aid of the relation of STUDIER.~~ For some esperiments native DNA was sonicated with a &lSE ultrasonic desintegrator. Thermal denaturation was achieved by heating DNA to IOO OC in 0.1 SSC (o-15 M NaCl + O.OI~ AI Na-citrate, pH 7) or o-ox SSC respecti\Tely in case of DNA4 from 3licrococcus Lysodeikfims and b>- subsequent rapid cooling in an ice bath. All measurements were made at 25 OC $0.1 OC in a thermostated METROHXI cell in which the solutions were adjusted at pH 5.6 with Mc ILV_AISE buffer and where the ionic strength was 0.5 M, adjusted by addition of KCl. Solutions were deaerated by passage of pure nitrogen for 30 minutes through the cell.

2. Sweep voltammetr~. - The measurements were carried out with a P_+R Polarograph Analyzer. Model 170. The built-in potentiostat and the electronic integrating circuit allowed the potentiostatic recording of current-potential curves and of charge-potential curves. The time constant was decreased by replacing the original condenser by one with about one order lower capacitance. For the recording of the cun-es a digital storage oscilloscope NICOLET, Model IOCJO.was used. The principles of the evaluation of the charge from the integrated cur\-es have been described previously.3 The working electrode was a METROHJI

hanging mercury drop electrode area of 3.50 mm”, the ausiliary reference

electrode

were

type E 410. with a surface

electrode was a platinum an TXGOLD saturated calomei electrode.

3. Alternating D.M.E.

(H.M.D.E.),

carried

coil and the

cnrrent polarography. - The measurement out with a P-AR P.olarographic Analyzer,

at

the

model

174. connected with a PAR Lock-in amplifier, model 122, and a PAR n-c. Polarographic Analyzer interface, model x74/30. The n.c. frequency selected was 78 Hz, the amplitude was adjusted to IO mV,,. The D.M.E. had the following characteristics : 112 = 0.23 mg s-r ; f, = 15.6 s at open

circuit in I Ad ICCL.

Results

and

discussion

The general measuring conditions for the sweep voltammetric studies are described for native calf thymus DNA as an esample in Fig. I and 2 where the dependence on the sweep rate (v) of the current

Table

I.

Sonica-

I Origin

DS_1

s t0.w *

ti0n

Keutral

Time

I

1

-1 CT2

I

I

15.6

3x106

/

( >fILES I

0

’

0

IS_'7

5-3 x 106

0

21.6

Sx106

SIG~lA

11.8

1 0-7x

106

I ;

I.SX

Ii

106

;

Labor.

CT3 CT,=

I

**

Labor.

IJO

S-2

23_s

; _#x

106

0.5

x IO6

21

7.4

X

IO6

19

f 2.7x106

gs

x

IO6

21.2

!

s

L

31%

?ttiLES

0

ML.,

Labor.

0

!

23.2

ML,b

Labor.

140s

1

7-s

I

x

0.45

IO6

3.2 x 106 -

I

ML4

SIGMA

0

I

2’2

gx

XIL,C

SIGMA

140s

i ’

IO

0-s x 106

CE,

i Labor_

0

21.6

sx

CEd

I Labor.

I

140s

;

9

0

:

23.2

IO6

106

20 -

[ 3 x IO6 .

-$x106

23

0.64 x 10~

I

PV,

i Labor.

g.s

x 106

I

P V,’

; Labor.

140s

I

9-f

Labor.

0

i

1g.s

i Labor.

140s

/

S.2

’

EC, EC,JPoiy-d(AT)

/

Polb--rh

- Polv-rU

j MILES

0

Poly-dG

- Poly-dC

i &IILES

0

I

i

0

.

bIILES

JIILES

0

0.52

17-7

: 2x106

16.4

;

x 106

6.2 x 10~

1.6x

x 106

-

-

10.2

0-g x IO6

-

-

1o.s

I

x 106

-

-

-

-

-

-

-

0.5

15.2 9-35

:0.2x106**=

ro6

Abbreviations used I CT : calf thymus DS_X : AIL : _~I~GYOCOGCICS LFsodeikticzrs DXA ; CE : chicken ecthrocyte DS_\ ; PV : Proteus T~‘zcZ~nris DSA ; EC : EscJterichia CoEi DSA. a _- starting from CT, ; b : starting from ML,, ; c : starting from ML, ; d ; starting from CE, ; CT.- starting from PV;; f : starting from EC,. Sedimentation coefficient (in Swedberg) corrected * Labor- corresponds to preparations from Centre Orleans, France. *** From Ref. 27_ *

l

for 20 oC_ de Biophysique

Mokulaire,

C.X.R.S.,

Electrochemical

Behaviour

of Satural

and

Biosynthetic

Polynecleotides

465

Fig. I. Sweep rate dependence of the redcction peak IP (Graph I) and its integrated value Q (Graph 2). Xative DNA CT,. ZOO pg/cm5, 0.5 M JICILVAISE buffer, pH 5.6. Adsorption time 70 s at an adsorption potential U, of - 0.4 V: H.M.D.E.

.

Fig. 2. Sweep rate dependence of the reduction peak IP (Graph I) and its inteo-rated value Q (Graph 2). Xative DNA CT?. zoo pg/cms, 0.3 AI >hdLV.UxE buffer, pH 5.6, Adsorption time 70 s at an adsorption potential U, of - 1.2 V. H.M.D.E.

p36

*

Sequaris,

JIalfoy.

valenta

and Xiirnberg

of the accessible II, and of the charge (Q) response due to the reduction reducible adenine and cytosine moieties of adsorbed polynucleotide is The reduction occurs in a total irreversible electrode process,” show-n. yielding a network of strongly adsorbed products blocking the electrode surface for adsorption of further DNA. 3 The measurements were carried out at the H.3I.D.E. at full coverage, the sweep starting and adsorption potentials rl; being -400 mV or -~200 mV. The time of adsorption (70 s) is long enough to allow the different interfacial events described stage. The dependence of in the previous paper S to reach a stationary the current (I,) on the sweep rate is linear in its first part* and the corresponding charge values [equal to the respective time integral of (IJJ remain independent of (u) over the whole range of the sweep rate (v). These results prove that reduction occurs only from adsorbed DNA under the esperimental conditions adjusted as usual (v = I V s-1 ; pH= The current or charge reduction responses are 5.6, AklL\-AISE buffer). nor are they not influenced by the diffusion of DN_1 from the solution controlled b>- the protonation kinetics of the reducible bases at the The charge is thus proportional to the amount of reducible adjusted pH. bases accessible to reduction in the adsorbed native DNA. The constancy of Q up to very high sweep rates (50 V s-l) suggested that, virtually only during the adsorption time elapsed at the respective adsorption potential U,. is a corresponding amount of regions of the adsorbed biopol>-mer transformed during a sequence of interfacial events from the double helical to the single stranded structure.These were discussed in detail in the preceeding paper.S This sequence of structural changes occurring cooperativelyat various regions of the adsorbed biopolymer will have a moderate o\-erall rate although it contains also fast steps occur-r-in g at certain potentials and proceeding rapidly for those moieties accessible to them. These rapid e\-ents in the sequence of interfacial structural rearrangements may therefore occur for those moieties which ha\-e become ready to undergo them during the sweep (for details see Ref. S). In general, the results shown in Fig. I and 2 confirm once more the fundamental feature3rr6*‘” of sweep and pulse methods : they both reflect mainly-, vin the reduction response, the interfacial situation which has developed at the sweep or pulse starting potential U, during the adsorption time elapsed there, provided that reduction is restricted to the adsorbed material ain the properly adjusted esperimental conditions. In Fig. 3 the dependence of the reduction response Q on the adsorption potential and sweep starting potential U, is shown for various doublestranded helical and single-stranded biopolymers differing in their content -4dsorption time was in all cases 70 s ; thus the of A-T paired bases. electrode surface was fully covered. The charge US. potential curves of the native DN_\ are always below the corresponding curves of the denatured

* Slight deviations from linearity at higher sweep rates are due to a slight remaining ohmic IR-cirop at ele\-ated sweep rates (3 > IO V s-l)_

Electrochemical

DNA. strand sponse totally

Behaviour

of

Natural

and

Biosynthetic

Polynucleotides

467

For the more negative CT-values, the unwinditig otthe doubleincreases, as described in previous papers.ls= The redtiction reof the DNA approaches the reduction charge obtain_tid:-with a : single-stranded form.

Fig. 3. Dependence of the reduction response Q of various DX-Xson adsor$ tion potential. 0.5 M MCILVAIXE buffer, pH 5.6, adsorption time 70 s. sweep rate I V/sI Xative ML, DNA ISO pg/cm*, z qenatured ML, DSX 50 pp”/crnz. 3 Xative CT, DNA 130 pg/crn5; 4 Denatured CT, DNA 50 ps/crnS, 5 Poly d(AT) 35 &cmS-

With respect_ to recent res&s reported by WEBB, JXNIK and ELon the steps of the electrode process of cytosine and adenink reduction from various oligonucleotides, the difkrences in the -reduction response of single-stranded bacterial and calf thymus DNA havingY_different contents of cytosine and adenine might find there their expk-. naticn. The somewhat increased Q-values at the most positive,. U,values observable for the bacterial native DNA as well as for both-kinds of denatured DNA is to be attributed -to some increase in coverage, due to electrostatic attraction, of the negatively charged phosphate soupS_? From the general pattern shown in Fig. 3, it will be noted that-the -various .. -: ’ ...*I !. charge responses depend on the relative content of adenine. In Fig. 4 the dependence of the..charge Q due to- the. reductionris plotted as a function of the adenine .percentage (% A) conkiined- in tl$e polynucleotides and. nucleic acids studied. AU Q-values -refer to th_e+ame sweep starting potential U, = -o.4 V and. thus- to the same_‘adsorpti& potential. They correspond to se,gment III, i.e. the plateau’ region; of- the dependence of the reduction response on the adsorption time af U, (cu&e The three biosynthetic double-stranded’ helical I in Fig. 3 of Ref. S). polynucleotides [ds-poly rA. ds-poly d(AT) tind ds-poly -rA-ply. .rIJj define a straight line running through zero. In these biosynthetic poly-‘::_ VISG~~

Sequaris.

Malfo?-.

Valenta

and

Xtirnberg

5-

”

s-2

0

3-

2-

l-

b 0

1

I 20

I

I 40

1

1 60

I

% Adenine 1 I 80

I 100

Fig. +_ Variation of the reduction response Q of various native DXAs and polynucleotides as a function of the percentage in adenine and of molecular weight 11~. CL5 11 JICILVAI~E buffer, pH 5.6, (pH 6 in case of single strand Poly rX) Sweep rate I V s-1 ; U, - 0.4 V. double single

strand

Poly

rX

25 &cms 23 pg/cmJ

strand Poly-r-1

Poly d(AT) Poly rX.Poly CTaDNX CT,DSA

(denatured)

CT,DNX dCT2DS_k bLL,DNA

ML,DNA

rU

35 wlcms 100 pg/cm5

S70 g&m3

Sg Fg/crna 150 &cm3 50 Erg/cm3 150 _vLglcms

50 ~g/cnl=

NW- : 0.2X106 Mw : 0.1 XL06 Mw : !,.a X106 JLW : I Xro’

Jiw : 0.5 x10= 3 X10’ 5.3Xx0’

NW : NW :

JIW : 9

XI06

>I\\~ : o.sxroe

nucleotides, the only reducible base is adenine, while DNA contains in its more stabile G-C-pairs also the reducible cytosine. The polynucleotides studied differ under various aspects in their structural details. Thus, the sugar moieties are deosyribose for poly-d(AT) and ribose for poly-rA-poly-rU and poly-rA. The double helis of the two polynucleotides, besides the general stabilizing hydration effects described by LEWIX,~ is maintained to a significant estent by WATSOSCRICK interstrand base pairin g hydrogen bonds as in DNA_ The double helix of poly-r‘x is maintained by interstrand hydrogen bridging between strands and also by interstrand only adenine-containin g complementary electrostatic interactions between protonated N(r) and negatively charged Of all the polynucleotides studied, poly-rA has .obphosphate groups. viously the least stable double helix Furthermore, as revealed by X-ray

Electrochemical

Behaviour

of Natural

and

Biosynthetic

Polynucle&ides

#3-j

diffraction for poly-rA-poly-rU,li poly-d(_ST)l* and ds-poly-rAl9 there exist differences between the respective polynucleotides with respect to the number of base pairs per turn of their helix, the dimension of their helis and the angle of the base pair plane with the helix axis. Nevertheless, the esperimental evidence in Fig. 3 shows that the differences in fine structure, that are observed in homogeneous phase, are actually insignificant at a charged interface where they are overruled by the effects due to the differences in adenine content and to the resulting consequences on double helis stability. For the case of IOO yO A represented by poly-rA, the Q-value, form (ss-poly-rA) indicates about 13 o/O larger for the single-stranded that, for the double-stranded form (ds-poly-rA), even in the stationary interfacial stage of unwinding,8 a certain amount of the reducible bases remains unaccessible to the electrode process. The electrode process and the interfacial behaviour of poly-rA has been recently clarified in detail behaviour was also studied by J;\XIK et by usRo The electrochemical nl. 21-23and by BRIBEC and P_+LECEK.“~~“~ Observ-ing carefully Fig. 4 once more, one realizes, from the results for native calf thymus DNX and for native bacterial DNA (Mic~ococcz~s Lysodeiktic,zls). that differences in the molecular weight of more than an Generally, the reduction order of magnitude esert also some influence. response Q decreases with increasin, m molecular weight for a native DNA with a given adenine content (% A). In this contest it should be noted that MALFOY and ~EYXXUD~ have recently shown by tensammetric measurements that the highest degree of adsorption is achieved for the lowest molecular weight of the respective type of native DNA. According to RECORD and WOODBURY,‘~ native DNA polymers with low molecular weight have a rod like shape, while at higher molecular weight values the double helix adopts progressively a coiled shape. Thus, the beforementioned authors stated that for a moiecular weight of I x IO”. the biopolymer in solution is to be extended to ST o/0of its contour length. For a molecular weight of 4 x 10~ the estension is only 30 o/o_At higher molecular weight, a coiled structure is progressively preferred. Therefore, the reduction response depends on the shape of the native DNA in solution. The influence of the macromolecule’s shape in the proposed model of unwinding is schematical!y described in Fig. 5_ .

Influence of the molecular u-eight on the behavior of native DNA at the electrode. _4 : high molecular weight ; B : low molecular weight.

470

Sequaris,

Xalfoy,

Valenta

and

Siirn berg

Concerning high molecular weight (Fig. 5 A), native DN_A has a coiIed structure in solution and adsorbs zin a large number of sites. At each contact point the unwinding is promoted in con\-ergent direction_ Some compact regions due to sterical constraints soon appear. The bases in those ‘regions are not accessible to reduction. For DN-1 of low molecular weight or sonicated DNA (Fig. 5 6) the number of the macromolecule’s contact points with the electrode is smaller. The promoted unwinding can now estend with Iess sterical macromolecule with onl>- one or constraint _ In Fig__ 5 a hypothetical fen- contact points IS schematically described. Considering the Influence eserted by the molecuIar weight upon the behal-iour of nati\*e DN-4. the comparison between the reduction responses of the various polynucleotides is only justified for polynucleotides of similar molecular weights. It is then possible to compare in Fig. _I the different charge values obtained with sonicated native DNA from dfr’crococczrs L. and calf thynlus and with the three Polk-nucleotides : poly-rX-poly-rU, poly-d(.;\T) , poly-r-4. The molecular weiiht of these compounds is somewhat smaller native DNA, than I s 10~. One notices that, with regard to the sonicated the charge values show no linear x-ariation with the percentage of adenkc

Fig. G. Vikiation of the potential T.IXI height of the n.c. peak (I) a~ ;I function of the perccntngc in A fT. Concentration of polyn~~cleotidcs : 400 .ugjcmJ. 0.3 :\I >ICIL\-AISE buffer. pH 3.6 : D.M.E. : drop time 7.6 s. e ML, D3.T.A; A EC, DS_A ; 0 P\-, DS_-\ ; 0 CT, DSA; n CE, DSA ; w polyd(.AT) (see table I).

EIectrochemicaI

Behaviour

of Satural

and

Biosynthetic

Polynucleotides

471

as it was observed with the three polynucleotides. This is due to the fact that in DNA there are also c>-tosine moieties involved in the adsorption at UI, of - 0.4 V and a subsequent reduction occurs during the sweep. The conclusions emerging from the relationship she\\-n in Fig. 4 of the skwep voltammetric reduction response Q on the adenine content (7.L 3) for the respective biopolymer are further confirmed by n.c.-polarographic measurements at the D.M.E. shown in Fig. 6. Here the height and potential of the capacitix-e n.c.-peak (out-of-phase component at go0 phase angle of the n.c. signal) due to the reorientation of the rotatable bases in the single-stranded stage of the adsorbed biopolymers has been studied as a function of the content of adenine + thymine [ Olo(X+-T)]. ,ks was described in detail in the preceding paper on time eftects,s this non-faradaic peak termed peak (I) [in Ref. S] is due to the orientation of the bases into a more perpendicular adsorption position with respect to the electrode surface in those regions of the biopolymer which have attained the single stranded stage. The height 1[ of the corresponding KC. peak grows linearly with increasing (A+T)-content of the respective nucleic acid and the peak potential r/; is shifted to more positive values also according to a linear dependence on y/o(X+T). For native DNA of different origin but w-ith the same (A+T)-percentage. virtually identical values of peak height and peak potential are obtained. The fact that the linear dependence of peak height on y. (-1+-T) runs nearly through the zero, permits the more detailed conclusion that the cc.c,-peak [i.e. peak (I) in Ref. S] is only related to the reorientation of adenine and thymine immediately after rupture of the WATSOX-CRICK bonds in the concomitant transinterstrand A-T-base pairin, (J hydrogen formation of the linear-stranded ladder structure into a single-stranded structure at these regions of the adsorbed biopoly-n~er.8 Considerations drawn from molecular models reveal that the reorientation of adenine and thymine (reflected by the capacitive n-c. peak treated in Fig. 6) from the interstrand base-paired stage in the ladder structure to the beforementioned position finally adopted in the single-stranded stage (after rupture of the* interstand base-pairing hydrogen bridge) is foregoing connected with significantly iarger local structural alterations involving the environmental sugar-phosphate moieties of the considered A-T-pairs than for a G-C-base pair in a similar sequence of local structural rearrangements. The results obtained for the capacitive n-c. response with the double helical forms of two biosynthetic polynucleotides (ds-poly-d(AT), dspoly-dG-poly-dC) which either contain only A-T or G-C base pairs’ fit also \-eq- well into the emerging picture. It respectively (Fig. 7) should be noted that the measurements are taken at the H.3I.D.E. after an adsorption time of only 7 s (to be compared with the drop life of 7-6 s of the D.3I.E. measurements in Fig. 6) at the respective adjusted value of the mean electrode potential I/‘,,,. Making the reasonable assumption of approsimately equal surface activity for both polynucleotides, the same and thus comparable degree of fractional coverage of the electrode surface will have been reached after 7 s, while the attainment of full

Sequaris,

Malfoy,

Valenta

and

Siirnberg

-

Poly d (AT)

-

Poly dG dC

l-

t -03

I

1 -05

I

I -0.7

I

I -09

I

, -1.1

I

U(V)

I

I

-1.3

-1.5

Fig. i_ Capacitive CL.~.compoucnt I, as a function of the adsorption potential U, (for detaik. see Ref.S) I JI NCILVAISE buffer, pH 5.6; Frequency 7s Hz. (IL. voltage 5 mVpp; adsorption time 7 s 1 ; Poiy d(AT) 25 Fg/cmJ; 2 Polg-dG.Pol!dC 50 Fg/cm3.

would require 65 s or more. The same reorientation cz.c. peak already discussed is obtained for ds-poly-cl(XT) (curve I in Fig. 7)_ Due to the onset of reduction at potentials U,,, more negative than -1.2 V, its negative side is blurred for this polynucleotide by the pseudocapacitance connected with the faradaic process. On the other hand, for ds-poly-dG-poly-dC only a shoulder is observable at somewhat more negative U,,,-values. thus becoming even more blurred by the onset of the It seems tempting to associate the observable shoulder with reduction. the visible part of a similar reorientation CI.C.response for the bases from the G-C-pairs bein,m somewhat smaller and appearing at more negative U,,,-values due to the greater stability and the smaller estent of local structural alterations discussed for G-C-pairs before. In conclusion, it appears that the high values of the reduction response obtained with r-n&e DNA imply that the structure of the DNA, as described by IVATSOK and CRICK, must be strongly modified for adsorbed DNA. The proposed model suggested by our results1a2.8 is an approach to the structure of the DNA at charged biological interfaces a location frequently adopted by DNA in living cells. coverage

Acknowledgement We are indebted to Prof. C. H~LI?:SE and Dr. G. SXDROX who have agreed to make the preparation, the sonication and the ultracentrifugation of DNA1 samples at the Centre de Biophysique Mokulaire, C.N.R.S.. Orleans, France.

Electrochemical

Behaviour

of Satural

and

Biosynthetic

and

NDRSBERG,

Biopltys.

Polynucleotides

473

References 1

I!

P. VXLESTA P. VXLESTX,

H.W.

H.W.

X~~RSBERG

and

P.

Stnrct.

Mech. 1, 17 (1974) Bioelecivochem. Bioeuer,o.

KLAIIRE,

1, -187 (1974) 3

P.

VALEX~A

them. .a

P.

49,

41

V.=.LESTA

and

P.

GRAI-IMAXX.

(1974) and H-X\'.

J_

Electroannl. J_

X~~RSBEZCG.

Chew.

Electvoa~zal.

49, 55 (1974) B. JANII~ and P. J. ELVISG, Cllem. Rev. 68, 295 B. MXLFOY and J__I. REYSXUD, J_ Electroaml.

EZectro-

Intevfacinl

Cilem.

Ixtevfacial

Elec-

frochenr. 5 6

them., 7 Y

9

10

11 12

13

67,

359

(196s) Chenr.

Inierfncinl

ELectvo-

(1976)

Collect. Csecll. Clrerlr. Co~w~~lwz. 39, 3-149 (1974) P. VXLESTX and H.\r. ~URSBERG, BioeiecB. MXLFO\-, J.M. SEQUARIS, tvochem. Bioeneq. see preceding paper S. LEN-IX. Displacenzext of IVnter ad its Control of Biochemical Reactions, Academic Press, Xew Sork (197-l) V. BRABEC and E. PXLE~EK. Z. Natwfovsch. Teil C 29, 323 (x97.+) J__\_ REX-SAUD, P.J. SICARD and A. OBRESOVITCH, Experientin 18, 543 (1971) J.-I. REX-SAL'D and P. J_ SICXRD, Bioelectrochem. Bioenevg. 1, B. AIALFOI-, 126 (1974) E.R.BI. KAY, J1.S. Sruuoss and -1.L. DOUXCE, J_ _+Irn. Cheer. Sot. 74, 127-1 E.

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19

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