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Pressure-induced spectral shifts in hemoproteins
mcnt
of Pl~srn~acology, Loriisiana State UniverPniniun sclerops from W. I?. Prince, Silver Springs, Florida. Synthet.ic I,-TIlCA was kitldly supplied ljy 1 jr. Thomas Briggs of the liniversity of Oklahoma. This work was sttpported in part by N.I.11. grants 05202.03 - .05. sily;
51. II.\sm\\.oo~), (;. A. I)., Niochr~~. J. 52, 583 (1952). 2. KL‘R.\KTI, Y., .\NI) K.~xuso, T., Hoppe-Seyl %. Phpiol. (‘hcnt 262, 53 (1939). 3. ~I.tHLTI, II., tf. I
Ikphys.
8. LLX,DY,
J. L., HERM.\N, &:., .\SD STAPLE,
R.
II.,
d?“Ch.
WEINf~idlH%
114, 233 (1966). F.
S(‘HZEIl)ER.
1:.,
ti.1RBORI), It.
w.,
Ii. J.
A., .tNI)
d WL. oil
1hXEN-
ChWZ.
SOC.
37, 447 il%iO). 1’. 1’. sII:\H
Veleran’s ~ldmini.sf~alion Hospiial Philadelphia, Pennayluania 1.9104 Receiucd Ikcc~mbcr 19, 1967
Pressure-Induced Spectral Hemoproteins
Shifts
in
We have observed the pressure-indticed spectral shift in the spectra of hemoglobin and it.s derivatives. The compouuds investigated were met,hemoglobin (pH 6.9), methemoglobin fltioride, methemoglobin cyanide, oxyhemoglobin, deoxyand hematoporphyrin hemoglobin, hemat,in, IX. Figttre 1 shows the rcsttlts for the SoretOband absorption. Within experimental error (f5 A) we saw no change of the posit.ion of the OLand fl band of oxyhemoglobin niid hematin or in the 560-rnp band of deoxghemoglohin up lo the pressure of 2000
atm.
The effects observed are an order of magnitude greater thati one would expect, by analogy to other a.romatic systems (1).
t f Jr i O- i -I -20 ’ ’ ’ ’ 1000I ’ ’ ’ ’ PRESSURE
2000
ATMOSPHERE
FIG. 1. Shift in the position of the Soret hand at 25’C for different hemoproteins. Hemoglobin was prepared from the freshly drawn blood of an adtilt male by washing the red blood cells with 0.9“; saline and centrifuging of the stroma after hemolysis. Concentrations were about lo-‘. Hemoglobin was oxidized with ferricyanide. Experiments were carried orit at pII 7. Ligand concentrations were 10-Z; phosphate bulfer eoncent.ration, IO+. Sigma Hematin was dissolved in 10m3 N NaO1-I. Hematoporphyrin IS 2 I-ICI (California Corp. for Biochemical Research) was dissolved in distilled water. A, Hematin; 0, oxyhemoglobiti; X, hematoporphyrin; 0, methemoglobin; 0, methemoglobin HLroride; n , methemoglobin cyanide. It, is well demonstrated that the position of the Soret band is proportional to the paramagnetic susceptibility of the methemoglobin compound (2). The advantages of investigating the high spinlow spin transition by presstrre, rather t.han by temperature variation, are twofold: (1) proteins are stable under pressures tip to 3000 atm, but they denattire under relatively low temperattire variations; (2) the Soret band positions in Fig. 1 cover the whole range from high to low spin forms, a rairge not, available by temperatrtre variations. All the absorption bands invesfigated are transition. According to preporphyrin r-r* vious observations (l), an approximately linear relationship can be obtained between wavelength shift tmder presstire and dielectric const,ant of the solvent. Since in the limited pressure range of our experiment, t,he dielectric constant of the solvent
Erratum Vol. 122, XhL)H and 501-50s : Page 503, Png;e 507,
-30. 2 (l!Xi), in the article entit’led, “Wurster’s Blue Alediated Oxidation of I’hosphorylation in ~Iitochondria,” hi- II. G. Mustafa and Two IS. King, pp. Icig. 1, ordinate should read: “FM Oz, SW.” Pig. ‘7, coupling site I ,should be located at, reaction step 2.
of Pl~srn~acology, Loriisiana State UniverPniniun sclerops from W. I?. Prince, Silver Springs, Florida. Synthet.ic I,-TIlCA was kitldly supplied ljy 1 jr. Thomas Briggs of the liniversity of Oklahoma. This work was sttpported in part by N.I.11. grants 05202.03 - .05. sily;
51. II.\sm\\.oo~), (;. A. I)., Niochr~~. J. 52, 583 (1952). 2. KL‘R.\KTI, Y., .\NI) K.~xuso, T., Hoppe-Seyl %. Phpiol. (‘hcnt 262, 53 (1939). 3. ~I.tHLTI, II., tf. I
Ikphys.
8. LLX,DY,
J. L., HERM.\N, &:., .\SD STAPLE,
R.
II.,
d?“Ch.
WEINf~idlH%
114, 233 (1966). F.
S(‘HZEIl)ER.
1:.,
ti.1RBORI), It.
w.,
Ii. J.
A., .tNI)
d WL. oil
1hXEN-
ChWZ.
SOC.
37, 447 il%iO). 1’. 1’. sII:\H
Veleran’s ~ldmini.sf~alion Hospiial Philadelphia, Pennayluania 1.9104 Receiucd Ikcc~mbcr 19, 1967
Pressure-Induced Spectral Hemoproteins
Shifts
in
We have observed the pressure-indticed spectral shift in the spectra of hemoglobin and it.s derivatives. The compouuds investigated were met,hemoglobin (pH 6.9), methemoglobin fltioride, methemoglobin cyanide, oxyhemoglobin, deoxyand hematoporphyrin hemoglobin, hemat,in, IX. Figttre 1 shows the rcsttlts for the SoretOband absorption. Within experimental error (f5 A) we saw no change of the posit.ion of the OLand fl band of oxyhemoglobin niid hematin or in the 560-rnp band of deoxghemoglohin up lo the pressure of 2000
atm.
The effects observed are an order of magnitude greater thati one would expect, by analogy to other a.romatic systems (1).
t f Jr i O- i -I -20 ’ ’ ’ ’ 1000I ’ ’ ’ ’ PRESSURE
2000
ATMOSPHERE
FIG. 1. Shift in the position of the Soret hand at 25’C for different hemoproteins. Hemoglobin was prepared from the freshly drawn blood of an adtilt male by washing the red blood cells with 0.9“; saline and centrifuging of the stroma after hemolysis. Concentrations were about lo-‘. Hemoglobin was oxidized with ferricyanide. Experiments were carried orit at pII 7. Ligand concentrations were 10-Z; phosphate bulfer eoncent.ration, IO+. Sigma Hematin was dissolved in 10m3 N NaO1-I. Hematoporphyrin IS 2 I-ICI (California Corp. for Biochemical Research) was dissolved in distilled water. A, Hematin; 0, oxyhemoglobiti; X, hematoporphyrin; 0, methemoglobin; 0, methemoglobin HLroride; n , methemoglobin cyanide. It, is well demonstrated that the position of the Soret band is proportional to the paramagnetic susceptibility of the methemoglobin compound (2). The advantages of investigating the high spinlow spin transition by presstrre, rather t.han by temperature variation, are twofold: (1) proteins are stable under pressures tip to 3000 atm, but they denattire under relatively low temperattire variations; (2) the Soret band positions in Fig. 1 cover the whole range from high to low spin forms, a rairge not, available by temperatrtre variations. All the absorption bands invesfigated are transition. According to preporphyrin r-r* vious observations (l), an approximately linear relationship can be obtained between wavelength shift tmder presstire and dielectric const,ant of the solvent. Since in the limited pressure range of our experiment, t,he dielectric constant of the solvent
Erratum Vol. 122, XhL)H and 501-50s : Page 503, Png;e 507,
-30. 2 (l!Xi), in the article entit’led, “Wurster’s Blue Alediated Oxidation of I’hosphorylation in ~Iitochondria,” hi- II. G. Mustafa and Two IS. King, pp. Icig. 1, ordinate should read: “FM Oz, SW.” Pig. ‘7, coupling site I ,should be located at, reaction step 2.