withiii expcriiiiciit;il u r o r over tlie raiige c)i extcii-. sions given in Fig. 1 is regarded as fortuitous. At extensioiis higher than cu. TOYd the collagcll libers evidently become t a u t and hence cause thc (J stress to rise abruptly with further extension. C o h cloublc bond chdracter. In fact, this plmaritp of' p i , beiiig a crL-stallinepolymer where elastic ~ n o d u the amide group should be expected to suppress lus and tensile strength are much higher tlian tliosc steric hindrance to rotation about the other bonds in oi elastin, prevents rupture of the fiber hulidlc ;tt tlie protein chain, thereby reducing the change of higher stresses. 111 contrast with rubber. tlic energy with chain conformation. Furthermore. steep rise in stress at. elongations near the iriasielastin is known16 to contain few polar amino inurn attainable leligth is not precipitated by crysacids and the electrical forces should not therefore tallization! but by- a permanently crystalline conibe expected to contribute appreciably to the con- Iionent (collagenj iiiterwol-en with the deforniable formational character of the chain. jelastinj component. Although the elastin bundle conforms to the conThe conclusions vie has-e reached depart from thc = 0 for ideal rubber elasticity dition ( b E b L ) v ~ interpretations of Ileyer and Ferri4 and of LYohlisch the linear stress-strain relation presents a striking and co-workers.5 Tliese investigators concluded contrast to the isotherm reported for rubber. This from stress- -temperature measurements that elastili feature finds qualitative explanation in the fact crystallizes on stretching. W6hlisch went even SO that elastin consists of separate fibers, curled to far :is to calculate the heat of crystallization o f clasdifferent degrees. Coiisequently some of the fibers tiii from the strcss--temper:tture relatioli. Thc ~ : i l u ~ . do not contribute a t all to the force a t low exteii- ioulld was surprisingly low. Failure to take acsioiis. .Is the extension iticreaies the proportioii of couiit of de-swelling of elastin in water with elevathe fibers subjected to extension increases; hence tion of temperature led these investigators to tltc more of them contribute to the force. The stress- erroiieous inferences. The internal energy changes strain relation which would obtain for a homogenc- with length as deduced by them are to be attributed o m polymer is therefore modified to the extent that rather to the eiiergy change associated with the the initial negative curvature which would other- riioreirientioric,d de-sw~ellinjiwith rise in ienilwraturc.. wise he obserwd is eliminatctl 'The 1iiic:irity noted
CC J A lit I t I I; I
1\
h I j . 1>iI;
PRI )hI L l I I:
s I l i I t [,I V I
1ti'. $1 1z 1'lt 1
I ,. IiI I JK.\ I I >I )
0.5 .5 .5 .a
.5 .5
~
D - D1 - __D2 - D I
1, 0 1.0 1. 0 1.0 1.0 5.0 5 .0 5.0 5 .0 5.0
Pco
Degree rif s a t n .
(cm.1
(%)
0.06 .19 .33 .66 .79 1.47 0.04 .09 .10 .16 .35 .05
42.6 68.4 79.7 88.0 90 . O 94.7 48.3 66.8 68 7 76.9 88.3 60.9 87.4 90.3 93.9 9i.3
,241 .24 .50 .97
experimental procedure was same as that given in Paper I. These results are summarized in Table I. Table I includes, in addition, data for heme solutions containing 5.0 X mole of pyridine per 1.
where D1 represents the optical density of one of the above solutions at a given wave length in the absence of carbon monoxide, D and DZ represent, respectively, the optical densities, a t the same wave length, of the same solution when it is partially and completely saturated with carbon monoxide. The validity of the abobc exprcssion for tlie dcgree of saturatioii X iiiay be shown as follows. Let f l i c i i t i ~ l ~ ct ru t i l i ( t i 1 1 1 1 ( I K liicit tit of 11.0 I I C I ~ I C - .iritl O~I~ 1) p> h C I l l ~c)IIz (b?cdLl\C i l f ~)bsLrvdlli111 e’ L tlic tnn1,tr c\tinctiori coc~lXciciitf J f II,O I~CIIIL C o atitl 115-henie-CO (bccduic o f u l ~ w r dtlc \ 11s 2 and 3 ) [Ii~O-hcme-OH~]= UI and [pj-lietne-OIl~] = bl at PLO= CJ P
1
[HzO-heme-CO] = c2 and [py-heme-CO] =d2 at satn. $ G O , [H2O-heme-OH2] = a 2o 10 [py-heme-OH*] , ,pco, , , , , , , Y O d t 5ome iiitermedi,tte ~HzO-heme-COl 0 [py-heme-CO] = d ~
Fig. 1.-
4 6 8 10 12 14 16 18 20 Partial pressure of CO (mm.). EITcrt of pyridine on the equilibrium between heme
2
6528
AKITSUGUNAKAIIARA AND
The spectrum of this solution in the absence of carbon monoxide is no longer the same as t h a t of solutions A, B and C but shows the presence of a minute amount of the complex py-heme-py. However the present procedure should still give the essentially correct value of the degree of saturation, provided t h a t the DI vaIue was determined from the corresponding absorption spectrum. The data in Table I, together with similar data in the absence of carbon monoxide reported in Paper I, are plotted in Fig. 1 for comparison. It is clearly shown in Fig. 1 that the small amounts of added pyridine markedly increase the affinity of the heme solutions for carbon monoxide. Since there is a reciprocal relationship between pyridine and carbon monoxide through their interaction with the same ferrous ion in heme, it may be inferred from above t h a t carbon monoxide also helps the heme to bind pyridine on the opposite side of the heme plane. This is not surprising if one recalls that when heme combines with carbon monoxide, the complex is changed from a high-spin to a low-spin state by the stronger total ligand-field of all six coordinating groups surrounding the ferrous ion in the new complex. The greater thermo-
JUI
Vol. 80
H. WANG
dynamic stability of py-heme-CO as coinpared to HzO-heme-CO is clearly due to the stronger ligand-field of the pyridine as compared to that of the water molecule. These heme complexes are particularly convenient models for ligand-field effect studies because of the absence of direct steric interference between the two coordinating groups on opposite sides of the heme plane. If the concentration of pyridine in a heme solution is increased continually, the afinity of the system for carbon monoxide should eventually decrease, because then the pyridine tends to combine with heme from both sides and compete with the carbon monoxide. It may also be of interest to note from Fig. 1 that the half-saturation pressure for carbon monoxide in 0.5 x lop4 M heme 5.0 X ill pyridine solution is 0.029 cm. which is only higher by a factor of 5 than the corresponding value for hemoglobin a t fiH 7.4. Acknowledgment.-This work was supported in part by a grant (USPHS-RG-4483) from the Division of Research Grants, u. S. Public Health Service.
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