The reaction of cysteine with ceruloplasmin copper

The kinetics of decay in absorbance at 610 nm in the reaction of cysteine with ceruloplasmin was biphasic under anaerobic conditions. Admission of oxygen to the bleached ceruloplasmin restored the blue color to about 75 % of the original value. However, under aerobic or anaerobic conditions an initial bleaching corresponded to a 25 % decrease in blue color. This change was irreversible and remained after removal of excess cysteine from the reaction mixture by dialysis. There was no correlation between transient and steady-state kinetic parameters. Circular dichroism measurements showed a characteristic reduction in the negative band at 450 nm, which is specific for type 1b copper. Isolation and further studies on cysteine-modified ceruloplasmin with a lower A₆₁₀/A₂₈₀ ratio showed < 10% reduction in enzyme activity toward p-phenylenediamine and o-dianisidine. Evidence is also presented that ceruloplasmin catalyzes the oxidation of cysteine with a one-electron reduction of oxygen and the formation of superoxide ion, which is then converted to H₂O₂ by ceruloplasmin. The effect of superoxide dismutase and catalase also confirms the presence of superoxide and H₂O₂. In sum, these data show that a permanent reduction of type 1b copper occurred when cysteine was used as a substrate. We conclude that there is a single electron transfer from cysteine directly to oxygen using one specific copper of ceruloplasmin, type 1b.     

The dimerization of protochlorophyll pigments in non-polar solvents

The infrared, visible and nuclear magnetic resonance spectra of protochlorophyll a and vinylprotochlorophyll a in dry non-polar solvents (carbon tetrachloride, chloroform, cyclohexane) are presented and interpreted in terms of dimer interaction.

The infrared spectra in the 1600–1800 cm⁻¹ region clearly show the existence of a coordination interaction between the C-9 ketone oxygen function of one molecule and the central magnesium atom of another molecule. Infrared spectra in the OH stretching region (3200–3800 cm⁻¹) provide a valuable test of the water content in the samples.

The analysis of the absorption and circular dichroism spectra of protochlorophyll a and vinylprotochlorophyll a in carbon tetrachloride demonstrates the existence of a monomer-dimer equilibrium in the concentration range from 10⁻⁶ to 5 · 10⁻⁴ M. The dimerization constants are (6±2) · 10⁵ 1 · M⁻¹ for protochlorophyll a and (4.5±2) · 10⁵ 1 · M⁻¹ for vinylprotochlorophyll a at 20 °C. The deconvolution of visible spectra in the red region has been performed in order to obtain quantitative information on the dimer structure. Two models involving a parallel or a perpendicular arrangement of the associated molecules are considered.

From ¹H NMR spectra, it appears that the region of overlap occurs near ring V, in agreement with the interpretation of the infrared spectra.     

Binding, internalization and intracellular processing of 125I-epidermal growth factor purified by isoelectric focusing

When epidermal growth factor (EGF) which had been extensively purified by HPLC was subjected to iodination with sodium ¹²⁵iodide, 5 major species of differing isoelectric points were produced. Some of these species bound to rat fibroblasts with different affinities but were internalized with equal efficiency. Examination of the internalized ¹²⁵I-labelled molecules revealed processing of all the ¹²⁵I-EGF species to macromolecules with more acidic isoelectric points. The ¹²⁵I-EGF species with a pI of 4.5 corresponded in electrofocusing behavior with intact non-iodinated EGF. Other EGF species probably represented molecules which were covalently modified as a result of the iodination procedure.     

Iron . bleomycin . DNA system evidence of a long-lived bleomycin . iron . oxygen intermediate

When Fe(II) is added to a bleomycin. DNA mixture in the presence of air a long-lived $\text{(}\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{= 45}\text{minutes}\text{)}$ EPR silent species (I′) is formed; the circular dichroism and absorption spectra of which have been characterized. This complex slowly decays yielding a ferric complex (III′) analogous to the well known low spin Fe(III). BLM species.     

Extrachromosomal human immunodeficiency virus type-1 DNA can initiate a spreading infection of HL-60 cells

In this report, we describe a human immunodeficiency virus type-1 (HIV-1)-infected promyelocytic cell line, OM, derived from HL-60 cells. Although the OM cell line was biologically cloned twice, the pattern of HIV-1 expression during culture appeared analogous to a classical acute spreading infection and was inhibited by both azidothymidine and recombinant soluble CD4 treatment. The number of OM cells actually expressing HIV-1 at the beginning of culture was 0%, reached a peak of nearly 100% at 6 weeks, and then fell to less than 10% HIV-1+ cells by 10 weeks. Clonal analysis of the surviving cells verified that stable HIV-1+ OM cells resulted from the spreading infection. Southern analysis confirmed the transmission of HIV-1 through these OM cultures and the occurrence of stable clones which resulted. The initial percentage of OM cells actually harboring the HIV-1 genome was less than 0.1%, indicating nonfaithful transmission of an unintegrated HIV-1 genome during clonal expansion. These results demonstrate that extrachromosomal HIV-1 DNA can contribute to the spread of HIV-1 infection and give rise to cells which have stably integrated HIV-1 provirus.     

Linear and circular dichroism of membranes from Rhodopseudomonas capsulata

Absorption, linear dichroism and circular dichroism spectra of Rhodopseudomonas capsulata (wild-type-St. Louis strain, mutant Y5 and mutant Ala⁺) are particularly sensitive to the nature of the light-harvesting bacteriochlorophyll-carotenoid-protein complexes. Evidence for exciton-type interactions is seen near 855 nm in the membranes from the wild-type and from mutant Y5, as well as in an isolated B-800 + 850 light-harvesting complex from mutant Y5. The strong circular dichroism that reflects these interactions is attenuated more than 10-fold in membranes from the Ala⁺ mutant, which lacks both B-800 + 850 and colored carotenoids and contains only the B-875 light-harvesting complex. These results lead to the conclusion that these two light-harvesting complexes have significantly different chromophore arrangements or local environments.     

Chemical modifications of actin: effects on interaction with deoxyribonuclease I

Maleylation of lysine residues, nitration of tyrosine residues or modification with 2,3-butanedione or 1,2-cyclohexanedione of arginine residues on actin resulted in a loss of polymerizability of the modified actin. However, only lysine modification produced a complete loss of the deoxyribunuclease I inhibitory ability of actin at low degrees of modification. By the level of one modified lysine per actin monomer, the samples completely lost polymerizability and lost 65% of their inhibitory power against deoxyribonuclease I-catalysed hydrolysis of DNA. By two lysines modified per actin, all inhibitory activity was lost. One lysine residue on actin apparently overlaps both an actin action contact site and an actin-deoxyribnuclease 1 contact site, offering a suggestion as to how deoxyribonuclease I blocks actin polymerization.