Biomedical subjects

J C Chien

Publications and source records attributed to J C Chien.

At least 37 records · Page 2Linked to original sources

PubMed1977

Electron paramagnetic resonance study of carp methemoglobin.

The g anisotropy of the EPR spectra of carp azidomethemoglobin is found to be pH-dependent, whereas, the spectra of human azidomethemoglobin are not. The two hemoglobins have the same g values at alkaline pH values. Crystal field analysis yielded values of 2.25 and 3.31, respectively, for the rhombic distortion, V/lambda, and the tetragonal distortion, delta/lambda. The spin orbit coupling constant is lambda. At pH 4.0 the values of V/lambda and delta/lambda for carp azidomethemoglobin became 1.95 and 4.76, respectively, whereas those for the human hemoglobin are virtually unchanged. The results are interpreted to mean an increase of out-ofplane displacement of the iron atom and stabilization of the T form of carp azidomethemoglobin by high proton concentration. At pH 6.0 and lower, the EPR spectra of carp azidomethemoglobin showed the presence of about 1.5% of high spin species, the amount is not affected by excess of either inositol hexaphosphate or sodium azide. The EPR spectra of aquo- and fluoroderivatives of carp methemoglobin were not affected by pH changes.

Animals↗

PubMed1977

Nonequivalence of subunits in [15N]nitrosylhemoglobin Kansas. A single crystal electron paramagnetic resonance investigation.

EPR spectra of Hb15NO crystals of mutant Kansas (Asn G4(102) beta leads to Thr) have been recorded at every 5' intervals and in three orthogonal planes. The nitrosylhemes are nonequivalent for the alpha and beta subunits, their assignments are made possible by comparison with the powder EPR specrtra of Hb15NO of mutant Iwate (His F8(87)alpha leads to Tyr) (Trittelvitz, E., Gersonde, K., and Winterhalter, K.H. (1975) Eur. J. Biochem. 51, 33-42). The EPR parameters for the beta-nitrosylhemes of Hb Kansas are: gxx=2.094 gyy=2.031, gzz=2.00, Azetazeta=11 G, Azetazeta=32.5 G, Aetaeta=12.5 G; the Fe-N-O bond angle is about 105 degrees. The paramters for the alpha-nitrosyl hemes are: gxx=2.058, gyy=2.021, gzz=1.977, Azetazeta=24.5 G, Azetazeta less than or equal to 5G, Aetaeta=23 G; the Fe-N-O bond angle is about 167 degrees. Hyperfine splittings of 7 to 8 gauss with 14Nepsilon atom of His(F8) were observed for the beta-nitrosylhemes; none was resolved for the alpha-nitrosylhemes. The results were interpreted to mean that the tension on the iron of the beta subunits is not large in the unliganded state and this tension was not greatly increased by the binding of nitric oxide in the strongly bent configuration. The tension at the iron in the deoxyhemoglobin is dominant at the alpha subunits. Binding of nitric oxide in this case causing either the breaking or great weakening of the Fe-His(F8) bond. The nitrosyl is in a nearly linear configuration. The unpaired electron densities at the nitrogen atom of the bound nitric oxide is about 63% for the beta-nitrosylheme and 37% for the alpha-nitrosylhemes.

Binding Sites↗

PubMed1976

Allosteric transitions in cobalt hemoglobins.

Circular dichroism and difference ultraviolet visible spectra were obtained for cobalt hemoglobin derivatives. At 287 nm the ellipticity difference between the oxy- and deoxycobaltohemoglobin is about one-half as great as that for the native proteins indicating smaller quaternary conformational changes for the former. Deoxygenation increases the Soret rotational strengths of both iron and cobalt hemoglobins to comparable degrees suggesting similar conformational changes for their aromatic residues near the "heme." Deoxygenation causes a much larger decrease of L band ellipticity for iron than cobalt hemoglobin. Circular dichroism spectra of nitrosylcobaltohemoglobin indicate the molecule to have a T quaternary structure. The circular dichroism spectra of cobaltihemoglobin do not seem to fit the patterns of the other cobalt derivatives and its 287 nm ellipticity is pH-dependent. From the shape of the Soret circular dichroism spectra, it is estimated that the transition dipole makes an angle with the line joining the two opposing pyrrole nitrogens of about 60 degrees for oxy- and deoxycobaltohemoglobin, 80 degrees for cobaltihemoglobin, as compared to 70 degrees for the native oxy- and deoxyhemoglobins. Inositol hexaphosphate has little or no effect on the circular dichroism spectra of cobalt hemoglobins in the 287 nm region, but it significantly increases the Soret rotational strength and decreases the L band ellipticity. The results are interpreted to mean that polyphosphates modify primarily the protein structure of hemoglobins at the tertiary level, and that the intersubunit interactions are weak in cobalt hemoglobins.

Binding Sites↗

PubMed1975

Cobalt-cytochrome c. I. Preparation, properties, and enzymic activity.

An improved procedure for the preparation of cobalt-cytochrome c has been developed. Various factors influencing the cobalt insertion process are discussed. The optical spectra of cobalt-cytochrome c suggest a six-coordinated species. The spectral shifts occurring with oxidation-reduction are compared with those observed for deoxy-cobaltohemoglobin and ferrocytochrome c and attributed to the effect of d(z2) electron on stereoelectronic interactions between the axial ligands and the porphyrin pi systems. Cobalt-cytochrome c has Em,7 = -140 +/- 20 mV as compared to an Em,7 of +250mV for ferrocytochrome c. An explanation for this negative Em,7 is offered. Cobaltocytochrome c is oxidized by cytochrome oxidase at about 45% of the rate for native cytochrome c. On the other hand cobalticytochrome c was not reduced by microsomal NADH or NADPH cytochrome c reductase nor by mitochondrial NADH or succinate cytochrome c reductase. It appears that the integrity of the reductase binding site is destroyed and the oxidase binding site has been modified by cobalt substitution.

Animals↗

PubMed1975

Cobalt-cytochrome c. II. Magnetic resonance spectra and conformational transitions.

Between pH approximately 4 and 10 cobaltocytochrome c (Cocyt-c) gives an electron paramagnetic resonance (EPR) spectrum with g parallel = 2.035, g the perpendicular = 2.223, CoA PARALLEL = 61.4 G, CoA the perpendicular = 49.8 G, NA parallel = 15.3 G, and NA THE PERPENDICULAR = 12.5 G. Comparisons with the EPR spectra of deoxycobaltomyoglobin, deoxycobaltohemoglobin, and model compounds and together with other evidence showed cobaltocytochrome c to have Met-80 and His-18 as its axial ligands. The protons of these ligands are seen as resonances shifted by the ring-current field of the porphyrin in the 300-MHZ 1H nuclear magnetic resonance (NMR) spectra of cobalticytochrome c (Cocyt-c+). The methyl and gamma-methylene protons of Met-80 in this molecule occupy positions with respect to heme c which are somewhat different from those in ferrocytochrome c. The 1H NMR spectra also showed that the methyl groups of Leu-32, Ile-75, Thr-63, thioether bridges, and the porphyrin ring in the cobalt protein are in the same state as in native enzyme; the same is also true for Tyr-59, His-26, and His-33 and also possibly Tyr-67, Tyr-74, and Phe-82. Above pH 11, Cocyt-c is converted to a five-coordinated form having g parallel = 2.026, g the perpendicular = 2.325, CoA parallel = 80 G, CoA the perpendicular approximately 10 G, NA parallel = 17.5 G, and NA the perpendicular not resolved. Below pH 1.0 the EPR spectrum of Cocyt-c is also five-coordinated with g parallel = 2.014, g the perpendicular = 2.359, CoA parallel = 93.8 G, and CoA the perpendicular = 38.8 G. The axial ligands in the alkaline and the acidic forms of Cocyt-c are His-18 and Met-80, respectively. New prominent proton resonance peaks are observed in cobalt-cytochrome c which are either absent or weak in native cytochrome c. These are situated at 3.0, 1.7, and 1.44 ppm, attributable, respectively, to the epsilon-CH2, DELTA-CH2 + beta-CH2, and gamma-CH2 of lysyl residues in random-coil-peptides. From the areas of these peaks, it is estimated that one-two lysyl residues in Cocyt-c have been modified; four-five lysyl residues in Cocyt-c+ have been modified. These alterations of surface charged groups are probably responsible for the lowered reactivity of Cocyt-c with cytochrome oxidase and the lack of reactivity of Cocyt-c+ with several cytochrome reductase systems.

Animals↗

PubMed1975

A 13C nuclear magnetic resonance and circular dichroism study of the collagen-gelatin transformation in enzyme solubilized collagen.

Natural abundance Fourier transform 13C nuclear magnetic resonance (13C NMR) were obtained for enzyme solubilized collagen at 1 degrees intervals through the transition region. The transition of collagen molecules from the rigid triple helical state to single-stranded, random-coil state is accompanied by a change from broadened carbon resonances unobservable under high-resolution conditions to narrow line spectra. Thus distinction can be made between helical and random-coil states of individual residues. The transition is monophasic, as determined by examination of 14 different carbon resonances, and the entire structure is found to melt cooperatively over a temperature interval of 5 +/- 1 degrees. All the residues seem to be involved in the unfolding process concurrently. The transition was also studied by examining the changes in the circular dichroism spectrum brought about by heating. The experiments corroborated the observation that the transition proceeded cooperatively over a temperature interval of 4 degrees. Enzyme soluble collagen is seen to melt less cooperatively than native collagen. The enthalpy change was determined by assuming an equilibrium between three random coil gelatin chains and tropocollogen molecules. From the enthalpy, the average length of the tripeptide sequences (70-85) involved in the transition can be estimated. The shortening of the cooperative unit could arise as a result of some alteration of the native conformation through proctase treatment.

Animals↗

PubMed1975